Synthesis and novel salt forms of (R)-5-((E)-2-pyrrolidin-3ylvinyl)pyrimidine

ABSTRACT

The present invention relates to the stereospecific synthesis of (R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine, its salt forms, and novel polymorphic forms of these salts.

FIELD OF THE INVENTION

The present invention relates to a stereospecific synthesis of(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine, its salt forms, and novelpolymorphic forms of these salts. The present invention also includespharmaceutical compositions of these salt forms as well as methods fortreating a wide variety of conditions and disorders, including pain,inflammation, and conditions and disorders associated with dysfunctionof the central and autonomic nervous systems.

BACKGROUND OF THE INVENTION

The compound (R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine is a neuronalnicotinic receptor (NNR) agonist with selectivity for the α4β2 nicotinicsubtype over other nicotinic subtypes, for example, the α7 subtype, theganglionic, and the muscle subtypes.(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine provides benefits in thetreatment or prevention of central nervous system (CNS) disorders andpain.

(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine has the followingstructural formula:

The commercial development of a drug candidate such as(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine involves many steps,including the development of a cost effective synthetic method that isadaptable to a large scale manufacturing process. Commercial developmentalso involves research regarding salt forms of the drug substance thatexhibit suitable purity, chemical stability, pharmaceutical properties,and characteristics that facilitate convenient handling and processing.Furthermore, compositions containing the drug substance should haveadequate shelf life. That is, they should not exhibit significantchanges in physicochemical characteristics such as, but not limited to,chemical composition, water content, density, hygroscopicity, andsolubility upon storage over an appreciable period of time.Additionally, reproducible and constant plasma concentration profiles ofdrug upon administration to a patient are also important factors.

Solid salt forms are generally preferred for oral formulations due totheir tendency to exhibit these properties in a preferential way; and inthe case of basic drugs such as(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine, acid addition salts areoften the preferred salt form. However, different salt forms varygreatly in their ability to impart these properties, and such propertiescannot be predicted with reasonable accuracy. For example, some saltsare solids at ambient temperatures, while other salts are liquids,viscous oils, or gums at ambient temperatures. Furthermore, some saltforms are stable to heat and light under extreme conditions and othersreadily decompose under much milder conditions. Thus, the development ofa suitable acid addition salt form of a basic drug for use in apharmaceutical composition is a highly unpredictable process.

The synthesis of 5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine and itshemi-galactarate salt, its separation by chiral chromatography intooptical isomers and the galatarate salts of the isomers are disclosed inpublished WO 04/078752 and U.S. Pat. No. 7,098,331, each of which isincorporated by reference. However, stereospecific syntheses of(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine, which are scalable to alarge-scale production, are desirable. Furthermore, because(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine in the free base form is aviscous oil with limited water solubility and stability, there is a needfor salt forms that display improved properties, including purity,stability, solubility, and bioavailability. Preferential characteristicsof these novel salt forms include those that would increase the ease orefficiency of manufacture of the active ingredient and its formulationinto a commercial product. Lastly, there is a need for stablepolymorphic forms of these salts that allows for an increase the ease orefficiency of manufacture of the active ingredient and its formulationinto a commercially product.

SUMMARY OF THE INVENTION

One aspect of the present invention is an acid addition salt of(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine. In certain embodiments,the acid is selected from hydrochloric, sulfuric, methanesulfonic,maleic, phosphoric, 1-hydroxy-2-naphthoic, ketoglutaric, malonic,L-tartaric, fumaric, citric, L-malic, hippuric, L-lactic, benzoic,succinic, adipic, acetic, nicotinic, propionic, orotic,4-hydroxybenzoic, di-p-toluoyl-D-tartaric, di-p-anisoyl-D-tartaric,di-benzoyl-D-tartaric, 10-camphorsulfonic, camphoric, or phencyphos.

One aspect of the invention is a maleic acid salt of(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine. Another aspect of theinvention is an orotic acid salt of(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine. A further aspect of theinvention is a citric acid salt of(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine.

One aspect of the invention is a(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine mono-citrate. Anotheraspect of the invention is a crystalline polymorph of(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine mono-citrate.

On aspect of the invention is a stereospecific synthesis of(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine. Other aspects andembodiments of the present invention will be described herein. The scopeof the present invention includes combinations of aspects, embodiments,and preferences.

BRIEF DESCRIPTION OF THE FIGURES

FIG. 1 is an XRPD pattern of amorphous form of(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine mono-citrate.

FIG. 2 is an XRPD pattern of Form I(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine mono-citrate.

FIG. 3 is an XRPD pattern of Form II(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine mono-citrate.

FIG. 4 is an XRPD pattern of Form III(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine mono-citrate.

FIG. 5 is an XRPD pattern of Form IV(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine mono-citrate.

FIG. 6 is an XRPD pattern of Form I(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine mono-orotate.

FIG. 7 is an XRPD pattern of Form I(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine mono-maleate.

FIG. 8 is an XRPD pattern of Form II(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine mono-maleate.

DETAILED DESCRIPTION

Definitions

The following definitions are meant to clarify, but not limit, the termsdefined. If a particular term used herein is not specifically defined,such term should not be considered indefinite. Rather, terms are usedwithin their accepted meanings.

The phrase “compounds of the present invention” as used herein refers to(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine or an acid addition saltthereof. The acid is selected from hydrochloric acid, sulfuric acid,methanesulfonic acid, maleic acid, phosphoric acid,1-hydroxy-2-naphthoic acid, ketoglutaric acid, malonic acid, L-tartaricacid, fumaric acid, citric acid, L-malic acid, hippuric acid, L-lacticacid, benzoic acid, succinic acid, adipic acid, acetic acid, nicotinicacid, propionic acid, orotic acid, 4-hydroxybenzoic acid,di-p-toluoyl-D-tartaric acid, di-p-anisoyl-D-tartaric acid,di-benzoyl-D-tartaric acid, 10-camphorsulfonic acid, camphoric acid, or2-hydroxy-5,5-dimethyl-4-phenyl-1,3,2-dioxaphosphorinan-2-one(phencyphos). The phrase includes a hydrate or a solvate form.

Further, as used herein, the term “compound” may be used to mean thefree base form, or alternatively, a salt form of(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine, depending on the context,which will be readily apparent. Those skilled in the art will be able todistinguish the difference.

As used herein, the term “pharmaceutically acceptable” refers tocarrier(s), diluent(s), excipient(s) or salt forms that are compatiblewith the other ingredients of the formulation and not deleterious to therecipient of the pharmaceutical composition.

As used herein, the term “pharmaceutical composition” refers to acompound of the present invention optionally admixed with one or morepharmaceutically acceptable carriers, diluents, excipients, oradjuvants. Pharmaceutical compositions preferably exhibit a degree ofstability to environmental conditions so as to make them suitable formanufacturing and commercialization purposes.

As used herein, the terms “effective amount,” “therapeutic amount,” or“effective dose” refer to an amount of active ingredient sufficient toelicit the desired pharmacological or therapeutic effects, thusresulting in effective prevention or treatment of a disorder. Preventionof a disorder may be manifested by delaying or preventing theprogression of the disorder, as well as delaying or preventing the onsetof the symptoms associated with the disorder. Treatment of the disordermay be manifested by a decrease or elimination of symptoms, inhibitionor reversal of the progression of the disorder, as well as any othercontribution to the well being of the patient.

The effective dose can vary, depending upon factors such as thecondition of the patient, the severity of the symptoms of the disorder,and the manner in which the pharmaceutical composition is administered.Typically, to be administered in an effective dose, compounds arerequired to be administered in an amount of less than 5 mg/kg of patientweight. Often, the compounds may be administered in an amount from lessthan about 1 mg/kg patient weight to less than about 100 μg/kg ofpatient weight, and occasionally between about 10 μg/kg to less than 100μg/kg of patient weight. The foregoing effective doses typicallyrepresent that amount administered as a single dose, or as one or moredoses administered over a 24 hours period. For human patients, theeffective dose of the compounds may require administering the compoundin an amount of at least about 1 mg/24 hr/patient, but not more thanabout 1000 mg/24 hr/patient, and often not more than about 500 mg/24 hr/patient.

As used herein, the phrase “substantially crystalline” includes greaterthan 20%, preferably greater than 30%, and more preferably greater than40% (e.g. greater than any of 50, 60, 70, 80, or 90%) crystalline.

The term “stability” as defined herein includes chemical stability andsolid state stability, where the phrase “chemical stability” includesthe potential to store salts of the invention in an isolated form, or inthe form of a formulation in which it is provided in admixture withpharmaceutically acceptable carriers, diluents, excipients, oradjuvants, such as in an oral dosage form, such as a tablet, capsule, orthe like, under normal storage conditions, with an insignificant degreeof chemical degradation or decomposition, and the phrase “solid statestability”, includes the potential to store salts of the invention in anisolated solid form, or in the form of a solid formulation in which itis provided in admixture with pharmaceutically acceptable carriers,diluents, excipients, or adjuvants, such as in an oral dosage form, suchas a tablet, capsule, or the like, under normal storage conditions, withan insignificant degree of solid state transformation, such ascrystallization, recrystallization, solid state phase transition,hydration, dehydration, solvation, or desolvation.

Examples of “normal storage conditions” include one or more oftemperatures of between −80° C. and 50° C., preferably between 0° C. and40° C. and more preferably ambient temperatures, such as 15° C. to 30°C., pressures of between 0.1 and 2 bars, preferably at atmosphericpressure, relative humidity of between 5 and 95%, preferably 10 to 60%,and exposure to 460 lux or less of UV/visible light, for prolongedperiods, such as greater than or equal to six months. Under suchconditions, salts of the invention may be found to be less than 5%, morepreferably less than 2%, and especially less than 1%, chemicallydegraded or decomposed, or solid state transformed, as appropriate. Theskilled person will appreciate that the above-mentioned upper and lowerlimits for temperature, pressure, and relative humidity representextremes of normal storage conditions, and that certain combinations ofthese extremes will not be experienced during normal storage (e.g. atemperature of 50° C. and a pressure of 0.1 bar).

Compounds

One embodiment of the present invention includes(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine (Formula I) or apharmaceutically acceptable salt thereof.

In one embodiment, the compound of Formula I or a pharmaceuticallyacceptable salt thereof is substantially pure. In one embodiment, thecompound of Formula I or a pharmaceutically acceptable salt thereof issubstantially free of (S)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine. Inone embodiment, the compound of Formula I or a pharmaceuticallyacceptable salt thereof is present in an amount of about 75% by weightcompared to (S)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine, preferablygreater than 85% by weight, more preferably greater than 95% by weight,more preferably greater than 98% by weight, and most preferably 99% byweight or greater.

One embodiment of the present invention includes a method for thepreparation of (R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine or apharmaceutically acceptable salt thereof containing less than 25%,preferably less than 15%, more preferably less than 5%, even morepreferably less than 2%, and most preferably less than 1% of(S)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine by weight. Anotherembodiment of the present invention includes a method for thepreparation of (R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine or apharmaceutically acceptable salt thereof containing less than 25%,preferably less than 15%, more preferable less than 5%, even morepreferably less than 2%, and most preferably less than 1% of(S)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine by weight, without the useof a chiral chromatographic separation step.

One embodiment of the present invention includes a method for thepreparation of (R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine or apharmaceutically acceptable salt thereof containing less than 25%,preferably less than 15%, more preferably less than 5%, even morepreferably less than 2%, and most preferably less than 1% of(S)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine by weight. Anotherembodiment of the present invention includes a method for thepreparation of (R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine or apharmaceutically acceptable salt thereof containing less than 25%,preferably less than 15%, more preferable less than 5%, even morepreferably less than 2%, and most preferably less than 1% of(S)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine by weight, without the useof a chiral chromatographic separation step. Thus, in one embodiment ofthe present invention, a method for the manufacture of substantiallypure (R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine is provided, withoutreliance upon chromatographic separation. One embodiment of the presentinvention includes a method of manufacturing a compound of the presentinvention on a commercial scale, namely where the method is fullyvalidated cGMP commercial scale active pharmaceutical ingredient (API)manufacturing, with reference to 21 CFR Parts 210 and 211, hereinincorporated by reference.

One embodiment of the present invention includes use of(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine or a pharmaceuticallyacceptable salt thereof in the manufacture of a medicament.

One embodiment of the present invention includes a method for thetreatment or prevention of a variety of disorders and dysfunctions,comprising administering to a mammal in need of such treatment, atherapeutically effective amount of(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine or a pharmaceuticallyacceptable salt thereof. More specifically, the disorder or dysfunctionmay be selected from the group consisting of CNS disorders,inflammation, inflammatory response associated with bacterial and/orviral infection, pain, metabolic syndrome, autoimmune disorders or otherdisorders described in further detail herein. Another embodiment of thepresent invention includes compounds that have utility as diagnosticagents and in receptor binding studies as described herein.

One embodiment of the present invention includes a pharmaceuticalcomposition comprising a therapeutically effective amount of(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine or a pharmaceuticallyacceptable salt thereof and one or more pharmaceutically acceptablecarrier. One embodiment of the present invention includes the use of apharmaceutical composition of the present invention in the manufactureof a medicament for treatment of central nervous system disorders anddysfunctions. Another embodiment of the present invention includes(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine or a pharmaceuticallyacceptable salt thereof with reference to any one of the Examples.Another embodiment of the present invention(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine or a pharmaceuticallyacceptable salt thereof for use as an active therapeutic substance.Another embodiment of the present invention includes(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine or a pharmaceuticallyacceptable salt thereof for use to modulate an NNR in a subject in needthereof. Another embodiment of the present invention includes(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine or a pharmaceuticallyacceptable salt thereof for use in the treatment or prevention ofconditions or disorders mediated by NNR. Another embodiment of thepresent invention includes a use(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine or a pharmaceuticallyacceptable salt thereof in the manufacture of a medicament for use ofmodulating NNR in a subject in need thereof.

Another embodiment of the present invention includes a use of(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine or a pharmaceuticallyacceptable salt thereof in the manufacture of a medicament for use inthe treatment or prevention of conditions or disorders mediated by NNR.Another embodiment of the present invention includes a method ofmodulating NNR in a subject in need thereof through the administrationof (R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine or a pharmaceuticallyacceptable salt thereof.

Unless otherwise stated, structures depicted herein are also meant toinclude compounds which differ only in the presence of one or moreisotopically enriched atoms. For example, compounds having the presentstructure except for the replacement of a hydrogen atom by deuterium ortritium, or the replacement of a carbon atom by ¹³C or ¹⁴C, or thereplacement of a nitrogen atom by ¹⁵N, or the replacement of an oxygenatom with ¹⁷O or ¹⁸O are within the scope of the invention. Suchisotopically labeled compounds are useful as research or diagnostictools.

As noted herein, the present invention includes specific representativecompounds, which are identified herein with particularity. The compoundsof this invention may be made by a variety of methods, includingwell-known standard synthetic methods. Illustrative general syntheticmethods are set out below and then specific compounds of the inventionare prepared in the working Examples.

In all of the examples described below, protecting groups for sensitiveor reactive groups are employed where necessary in accordance withgeneral principles of synthetic chemistry. Protecting groups aremanipulated according to standard methods of organic synthesis (T. W.Green and P. G. M. Wuts, Protecting Groups in Organic Synthesis, 3^(rd)Edition, John Wiley & Sons, New York (1999)). These groups are removedat a convenient stage of the compound synthesis using methods that arereadily apparent to those skilled in the art. The selection of processesas well as the reaction conditions and order of their execution shall beconsistent with the preparation of compounds of the present invention.

The present invention also provides a method for the synthesis ofcompounds useful as intermediates.

General Synthetic Methods

One aspect of the present invention includes the method for thestereospecific synthesis of (R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine(11) outlined in Scheme 1. Commercially available tert-butyl(R)-3-hydroxypyrrolidine-1-carboxylate (compound 1) is treated withmethanesulfonyl chloride to give tert-butyl(R)-3-(methylsulfonyloxy)pyrrolidine-1-carboxylate (compound 2), whichthen is reacted with diethylmalonate and a suitable base (e.g.,potassium tert-butoxide or sodium ethoxide) to give diethyl(R)-2-(1-(tert-butoxycarbonyl)pyrrolidin-3-yl)malonate (compound 3) withinverted stereochemistry around the chiral carbon.

Suitable solvents for these reactions may be selected from the group oftoluene, xylenes,1-methyl-2-pyrrolidinone, dimethylformamide,dimethylacetamide, ethanol, tert-butanol, tetrahydrofuran,1,2-dimethoxyethane, dioxane, and mixtures thereof. In one embodimentthe solvent for the methanesulfonic ester formation toluene, and thesolvent for the malonate displacement is 1-methyl-2-pyrrolidinone. Inanother embodiment the solvent for the malonate displacement is ethanol.Suitable bases for these reactions may be selected from the group oftriethylamine, diethylisopropylamine, diisopropylethylamine, potassiumtert-butoxide, sodium metal, sodium hydride, sodium ethoxide, potassiumhydride and lithium hydride. In one embodiment the base for themethanesulfonic ester formation is triethylamine, and the base for themalonate displacement is potassium tert-butoxide. In another embodimentthe base for the malonate displacement is sodium ethoxide.

Hydrolysis of diester 3 with aqueous potassium hydroxide yields(R)-2-(1-(tert-butoxycarbonyl)pyrrolidin-3-yl)malonic acid (compound 4),which is decarboxylated to afford(R)-2-(1-(tert-butoxycarbonyl)pyrrolidin-3-yl)acetic acid (compound 5).Suitable solvents for these reactions may be selected from the group ofwater, ethanol, tetrahydrofuran, dimethylformamide, dimethylacetamide,1,2-dimethoxyethane, dioxane, 1-methyl-2-pyrrolidinone, toluene,dimethylsulfoxide, and mixtures thereof. In one embodiment the solventfor the ester hydrolysis is aqueous tetrahydrofuran, and the solvent forthe decarboxylation is 1-methyl-2-pyrrolidinone. In another embodimentthe solvent for the ester hydrolysis is ethanol, and the solvent for thedecarboxylation is a mixture of dimethylsufloxide and toluene. Suitablebases for the hydrolysis reaction may be selected from the group ofpotassium hydroxide, sodium hydroxide, potassium carbonate, sodiumcarbonate, barium hydroxide and cesium carbonate. In one embodiment thebase is potassium hydroxide. Reduction of compound 5 gives tert-butyl(R)-3-(2-hydroxyethyl)pyrrolidine-1-carboxylate (compound 6), which maybe reacted with methanesulfonyl chloride and then sodium iodide to givetert-butyl (R)-3-(2-(methylsulfonyloxy)ethyl)pyrrolidine-1-carboxylate(compound 7) and tert-butyl (R)-3-(2-iodoethyl)pyrrolidine-1-carboxylate(compound 8), respectively. Suitable solvents for the reduction reactionmay be selected from the group of tetrahydrofuran, ether, dioxane,1,2-dimethoxyethane, and mixtures thereof. In one embodiment the solventis tetrahydrofuran. Suitable reducing agents may be selected from thegroup of borane, diborane, borane-tetrahydrofuran complex,borane-dimethyl ether complex and borane-dimethylsulfide complex.Suitable solvents for the methanesulfonic ester formation may beselected from the group of toluene, xylenes, ether, tetrahydrofuran,1,2-dimethoxyethane, dioxane, and mixtures thereof. In one embodimentthe solvent for the methanesulfonic ester formation is toluene. Suitablebases for the methanesulfonic ester formation may be selected from thegroup of triethylamine, diethylisopropylamine and diisopropylethylamine.In one embodiment the base for the methanesulfonic ester formation istriethylamine. Suitable solvents for the iodide displacement may beselected from the group of 1-methyl-2-pyrrolidinone, dimethylformamide,dimethylacetamide, ethanol, tert-butanol, tetrahydrofuran,1,2-dimethoxyethane, dioxane, dimethylsulfoxide, and mixtures thereof.In one embodiment the solvent for the iodide displacement is1,2-dimethoxyethane.

Finally, treatment of compound 8 with potassium tert-butoxide gives ofcompound 9. Suitable solvents for this reaction may be selected from thegroup of 1,2-dimethoxyethane, 1-methyl-2-pyrrolidinone,dimethylformamide, dimethylacetamide, ethanol, tetrahydrofuran, dioxaneand mixtures thereof. In one embodiment the solvent is1,2-dimethoxyethane. Suitable bases for this reaction may be selectedfrom the group of potassium tert-butoxide, sodium ethoxide anddiazabicycloundecane. In another embodiment the base is potassiumtert-butoxide.

Palladium-catalyzed coupling of compound 9 with 5-bromopyrimidine yields(R)-1-(tert-butoxycarbonyl)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine(10), which is de-protected in the final step to give(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine (11). Suitable solvents forthe palladium-catalyzed coupling reaction may be selected from the groupof 1-methyl-2-pyrrolidinone, dimethylformamide, dimethylacetamide andacetonitrile. In one embodiment the solvent is dimethylacetamide.Suitable bases for the palladium catalyzed coupling reaction may beselected from the group of triethylamine, diethylisopropylamine,diisopropylethylamine, and sodium acetate. In one embodiment the base issodium acetate. Suitable phosphine ligands for the palladium catalyzedcoupling reaction may be selected from the group oftri-n-butylphosphine, tri-tert-butylphosphine, tricyclohexylphosphine,triphenylphosphine, tri-o-tolylphosphine and 1,1′-bis(diphenylphosphino)ferrocene. In one embodiment the phosphineligand is 1, 1′-bis(diphenylphosphino)ferrocene. Suitable palladiumcatalysts for the palladium catalyzed coupling reaction may be selectedfrom the group of palladium acetate, palladium chloride and dipalladiumtris(dibenzylacetone). In one embodiment the palladium catalyst ispalladium acetate. Suitable solvents for the de-protection reaction maybe selected from the group of water, dichloromethane, chloroform anddichloroethane. In one embodiment the solvent is water. Suitable acidsfor the de-protection reaction may be selected from the group oftrifluoroacetic acid, hydrochloric acid and sulfuric acid. In oneembodiment the acid is hydrochloric acid.

Those skilled in the art of organic synthesis will appreciate that thereexist multiple means of producing compounds of the present inventionwhich are labeled with a radioisotope appropriate to various diagnosticuses. For example, coupling of ¹¹C-labeled 5-bromopyrimidine withcompound 9 or followed by removal of the protecting group as describedwill produce a compound suitable for use in positron emissiontomography.

Salt Forms

One aspect of the present invention relates to novel salt forms of(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine.(R)-5-((E)-2-Pyrrolidin-3-ylvinyl)pyrimidine in the free base form is aviscous oil with limited water solubility. However, the free base willreact with both inorganic and organic acids to make certain acidaddition salts that have physical properties that are advantageous forthe preparation of pharmaceutical compositions such as crystallinity,water solubility, and stability toward chemical degradation. Typicallythese salt forms are pharmaceutically acceptable salts. One aspect ofthe present invention includes acid addition salts of(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine. The acid is selected fromhydrochloric acid, sulfuric acid, methanesulfonic acid, maleic acid,phosphoric acid, 1-hydroxy-2-naphthoic acid, ketoglutaric acid, malonicacid, L-tartaric acid, fumaric acid, citric acid, L-malic acid, hippuricacid, L-lactic acid, benzoic acid, succinic acid, adipic acid, aceticacid, nicotinic acid, propionic acid, orotic acid, 4-hydroxybenzoicacid, di-p-toluoyl-D-tartaric acid, di-p-anisoyl-D-tartaric acid,di-benzoyl-D-tartaric acid, 10-camphorsulfonic acid, camphoric acid, andphencyphos. The present invention also includes hydrates and solvates ofthese salt forms.

The stoichiometry of the salts comprising the present invention canvary. For example, it is typical that the molar ratio of acid to(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine is 1:2 or 1:1, but otherratios, such as 3:1, 1:3, 2:3, 3:2 and 2:1, are possible. Depending uponthe manner by which the salts described herein are formed, the salts canhave crystal structures that occlude solvents that are present duringsalt formation. Thus, the salts can occur as hydrates and other solvatesof varying stoichiometry of solvent relative to(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine.

In one embodiment of the present invention, the salt has a stoichiometryof acid to (R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine of 1:2. Inanother embodiment, the salt has a stoichiometry of acid to(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine of 1:1.

Another embodiment of the present invention includes(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine mono-citrate or a hydrateor solvate thereof. Another embodiment of the present invention includes(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine mono-orotate or a hydrateor solvate thereof. Another embodiment of the present invention includes(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine mono-maleate or a hydrateor solvate thereof.

A further aspect of the present invention comprises processes for thepreparation of the salts. The precise conditions under which the saltsare formed may be empirically determined. The salts may be obtained bycrystallization under controlled conditions.

The method for preparing the salt forms can vary. The preparation of(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine salt forms typicallyinvolves:

-   (i) mixing the free base, or a solution of the free base of suitably    pure (R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine in a suitable    solvent, with any of the acids in pure form or as a solution of any    of the acids in a suitable solvent, typically 0.5 to 1 equivalents    of the acid,-   (ii) (a) cooling the resulting salt solution if necessary to cause    precipitation, or-   (ii) (b) adding a suitable anti-solvent to cause precipitation, or-   (ii) (c) evaporating the first solvent and adding and new solvent    and repeating either steps (ii) (a) or step (ii) (b), and-   (iii) filtering to collect the salt, and optional recrystallization.

The stoichiometry, solvent mix, solute concentration, and temperatureemployed can vary. Representative solvents that can be used to prepareor recrystallize the salt forms include, without limitation, ethanol,methanol, isopropyl alcohol, isopropyl acetate, acetone, ethyl acetate,toluene, water, methyl ethyl ketone, methyl isobutyl ketone, tert-butylmethyl ether, tetrahydrofuran, dichloromethane, n-heptane, andacetonitrile.

One embodiment of the present invention comprises the hydrochloric acid,sulfuric acid, methanesulfonic acid, maleic acid, phosphoric acid,1-hydroxy-2-naphthoic acid, ketoglutaric acid, malonic acid, L-tartaricacid, fumaric acid, citric acid, L-malic acid, hippuric acid, L-lacticacid, benzoic acid, succinic acid, adipic acid, acetic acid, nicotinicacid, propionic acid, orotic acid, 4-hydroxybenzoic acid,di-p-toluoyl-D-tartaric acid, di-p-anisoyl-D-tartaric acid,di-benzoyl-D-tartaric acid, 10-camphorsulfonic acid, camphoric acid, andphencyphos salts of (R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine insubstantially crystalline form.

The degree (%) of crystallinity may be determined by the skilled personusing x-ray powder diffraction (XRPD). Other techniques, such as solidstate NMR, FT-IR, Raman spectroscopy, differential scanning calorimetry(DSC) and microcalorimetry, may also be used. For compounds of thecurrent invention, it has been found to be possible to produce salts informs which are greater than 80% crystalline.

Several of these crystalline salts demonstrated stability sufficient toestablish their promise in the production of pharmaceuticalpreparations. Such stability can be demonstrated in a variety of ways.Propensity to gain and release atmospheric moisture can be assessed bydynamic vapor sorption (DVS). Stability to elevated temperatures andhumidity can be studied by storing the solid salts at 40° C./75% RH forup to eight days, and then re-examining each by weight, appearance underthe microscope, and XRPD.

Polymorphs

The compounds of the present invention may crystallize in more than oneform, a characteristic known as polymorphism, and such polymorphic forms(“polymorphs”) are within the scope of the present invention.Polymorphism generally can occur as a response to changes intemperature, pressure, or both. Polymorphism can also result fromvariations in the crystallization process. Polymorphs can bedistinguished by various physical characteristics known in the art suchas XRPD patterns (diffractograms), solubility in various solvents, andmelting point.

The present invention includes various polymorphic forms of the saltforms of (R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine, includinghydrates and solvates of the salts. Such polymorphic forms arecharacterized by their x-ray powder diffraction (XRPD) patterns(diffractograms).

One embodiment of the present invention includes a crystalline form of(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine mono-citrate. Anotherembodiment of the present invention includes an amorphous form of(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine mono-citrate. Anotherembodiment of the present invention includes an amorphous form of(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine mono-citrate whose XRPDpattern substantially corresponds to that shown in FIG. 1.

One embodiment of the present invention includes a polymorphic form of(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine mono-citrate Form Icharacterized by a XRPD pattern comprising at least one of the followingpeaks:

2θ  5.27 10.03 13.77 21.73

Another embodiment, the present invention includes a polymorphic form of(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine mono-citrate Form I whoseXRPD pattern substantially corresponds to that shown in FIG. 2.

One embodiment of the present invention includes a polymorphic form of(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine mono-citrate Form IIcharacterized by a powder x-ray diffraction pattern comprising at leastone of the following peaks:

2θ 11.02 20.01 22.06 24.66 32.13 33.35 34.61 35.96 38.65 40.23

Another embodiment, the present invention includes a polymorphic form of(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine mono-citrate Form II whoseXRPD pattern substantially corresponds to that shown in FIG. 3.

One embodiment of the present invention includes a polymorphic form of(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine mono-citrate Form IIIcharacterized by a XRPD pattern comprising at least one of the followingpeaks:

2θ  9.43 12.24 16.24 18.38 19.18 19.48 21.52 22.89 23.08 24.28 30.7731.27 32.36 33.09 34.86 37.26 37.63 39.47

Another embodiment, the present invention includes a polymorphic form of(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine mono-citrate Form III whoseXRPD pattern substantially corresponds to that shown in FIG. 4.

One embodiment of the present invention includes a polymorphic form of(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine mono-citrate Form IVcharacterized by a XRPD pattern comprising at least one of the followingpeaks:

2θ  5.05 10.81 14.06 15.20 17.43 23.57 24.21 25.52 26.95

Another embodiment, the present invention includes a polymorphic form of(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine mono-citrate Form IV whoseXRPD pattern substantially corresponds to that shown in FIG. 5

One embodiment of the present invention includes a crystalline form of(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine mono-orotate.

One embodiment of the present invention includes a polymorphic form of(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine mono-orotate Form Icharacterized by a XRPD pattern comprising at least one of the followingpeaks:

2θ 2.55 6.54 8.66 13.26 14.56 15.98 17.47 18.53 19.30 20.26 21.05 22.0223.14 24.32 25.56 26.87 27.84 28.76 29.53

Another embodiment, the present invention includes a polymorphic form of(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine mono-orotate Form I whoseXRPD pattern substantially corresponds to that shown in FIG. 6.

One embodiment of the present invention includes a crystalline form of(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine mono-maleate.

One embodiment of the present invention includes a polymorphic form of(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine mono-maleate Form Icharacterized by a XRPD pattern comprising at least one of the followingpeaks:

2θ 12.81 16.09 18.00 19.07 24.49 26.40 26.04 27.88

Another embodiment, the present invention includes a polymorphic form of(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine mono-maleate Form I whoseXRPD pattern substantially corresponds to that shown in FIG. 7.

One embodiment of the present invention includes a polymorphic form of(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine mono-maleate Form IIcharacterized by a XRPD pattern comprising at least one of the followingpeaks:

2θ  4.31 16.56 18.29 18.78 19.64 20.27 21.02 21.46 21.90 22.43 22.8625.40 25.73 26.15 26.56 27.40 28.59 29.57

Another embodiment, the present invention includes a polymorphic form of(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine mono-maleate Form II whoseXRPD pattern substantially corresponds to that shown in FIG. 8

As noted, the salt forms of (R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidinemay exist in solvated, for example hydrated, as well as unsolvatedforms. The present invention encompasses all such forms.

The present invention also includes isotopically labeled compoundswherein one or more atoms are replaced by an atom having an atomic massor mass number different from the atomic mass or mass number usuallyfound in nature. Examples of isotopes that can be incorporated intocompounds of the invention include isotopes of hydrogen, carbon,nitrogen, oxygen, phosphorous, sulfur, fluorine, and chlorine, such as²H, ³H, ¹³C, ¹⁴C, ¹⁵N, ¹⁸O, and ¹⁷O. Such isotopically labeled compoundsare useful as research or diagnostic tools.

Pharmaceutical Compositions

Although it is possible to administer the compound of the presentinvention in the form of a bulk active chemical, it is preferred toadminister the compound in the form of a pharmaceutical composition orformulation. Thus, one aspect the present invention includespharmaceutical compositions comprising the compound of the presentinvention and one or more pharmaceutically acceptable carriers,diluents, or excipients. Another aspect of the invention provides aprocess for the preparation of a pharmaceutical composition, includingadmixing the compound of the present invention with one or morepharmaceutically acceptable carriers, diluents or excipients.

The manner in which the compound of the present invention isadministered can vary. The compound of the present invention ispreferably administered orally. Preferred pharmaceutical compositionsfor oral administration include tablets, capsules, caplets, syrups,solutions, and suspensions. The pharmaceutical compositions of thepresent invention may be provided in modified release dosage forms suchas time-release tablet and capsule formulations.

The pharmaceutical compositions can also be administered via injection,namely, intravenously, intramuscularly, subcutaneously,intraperitoneally, intraarterially, intrathecally, andintracerebroventricularly. Intravenous administration is a preferredmethod of injection. Suitable carriers for injection are well known tothose of skill in the art and include 5% dextrose solutions, saline, andphosphate buffered saline.

The formulations may also be administered using other means, forexample, rectal administration. Formulations useful for rectaladministration, such as suppositories, are well known to those of skillin the art. The compounds can also be administered by inhalation, forexample, in the form of an aerosol; topically, such as, in lotion form;transdermally, such as, using a transdermal patch (for example, by usingtechnology that is commercially available from Novartis and AlzaCorporation), by powder injection, or by buccal, sublingual, orintranasal absorption.

Pharmaceutical compositions may be formulated in unit dose form, or inmultiple or subunit doses

The administration of the pharmaceutical compositions described hereincan be intermittent, or at a gradual, continuous, constant or controlledrate. The pharmaceutical compositions may be administered to awarm-blooded animal, for example, a mammal such as a mouse, rat, cat,rabbit, dog, pig, cow, or monkey; but advantageously is administered toa human being. In addition, the time of day and the number of times perday that the pharmaceutical composition is administered can vary.

The compound of the present invention may be used in the treatment of avariety of disorders and conditions and, as such, may be used incombination with a variety of other suitable therapeutic agents usefulin the treatment or prophylaxis of those disorders or conditions. Thus,one embodiment of the present invention includes the administration ofthe compound of the present invention in combination with othertherapeutic compounds. For example, the compound of the presentinvention can be used in combination with other NNR ligands (such asvarenicline), antioxidants (such as free radical scavenging agents),antibacterial agents (such as penicillin antibiotics), antiviral agents(such as nucleoside analogs, like zidovudine and acyclovir),anticoagulants (such as warfarin), anti-inflammatory agents (such asNSAIDs), anti-pyretics, analgesics, anesthetics (such as used insurgery), acetylcholinesterase inhibitors (such as donepezil andgalantamine), antipsychotics (such as haloperidol, clozapine,olanzapine, and quetiapine), immuno-suppressants (such as cyclosporinand methotrexate), neuroprotective agents, steroids (such as steroidhormones), corticosteroids (such as dexamethasone, predisone, andhydrocortisone), vitamins, minerals, nutraceuticals, anti-depressants(such as imipramine, fluoxetine, paroxetine, escitalopram, sertraline,venlafaxine, and duloxetine), anxiolytics (such as alprazolam andbuspirone), anticonvulsants (such as phenytoin and gabapentin),vasodilators (such as prazosin and sildenafil), mood stabilizers (suchas valproate and aripiprazole), anti-cancer drugs (such asanti-proliferatives), antihypertensive agents (such as atenolol,clonidine, amlopidine, verapamil, and olmesartan), laxatives, stoolsofteners, diuretics (such as furosemide), anti-spasmotics (such asdicyclomine), anti-dyskinetic agents, and anti-ulcer medications (suchas esomeprazole). Such a combination of pharmaceutically active agentsmay be administered together or separately and, when administeredseparately, administration may occur simultaneously or sequentially, inany order. The amounts of the compounds or agents and the relativetimings of administration will be selected in order to achieve thedesired therapeutic effect. The administration in combination of acompound of the present invention with other treatment agents may be incombination by administration concomitantly in: (1) a unitarypharmaceutical composition including both compounds, or (2) separatepharmaceutical compositions each including one of the compounds.Alternatively, the combination may be administered separately in asequential manner wherein one treatment agent is administered first andthe other second. Such sequential administration may be close in time orremote in time.

Another aspect of the present invention includes combination therapycomprising administering to the subject a therapeutically orprophylactically effective amount of the compound of the presentinvention and one or more other therapy including chemotherapy,radiation therapy, gene therapy, or immunotherapy.

Method of Treatment

(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine, a pharmaceuticallyacceptable salt thereof, or a pharmaceutical composition containing suchcan be used for the prevention or treatment of various conditions ordisorders for which other types of nicotinic compounds have beenproposed or are shown to be useful as therapeutics, such as CNSdisorders, inflammation, inflammatory response associated with bacterialand/or viral infection, pain, metabolic syndrome, autoimmune disordersor other disorders described in further detail herein. This compound canalso be used as a diagnostic agent in receptor binding studies (in vitroand in vivo). Such therapeutic and other teachings are described, forexample, in references previously listed herein, including Williams etal., Drug News Perspec. 7(4): 205 (1994), Americ et al., CNS Drug Rev.1(1): 1-26 (1995), Arneric et al., Exp. Opin. Invest. Drugs 5(1): 79-100(1996), Bencherif et al., J. Pharmacol. Exp. Ther. 279: 1413 (1996),Lippiello et al., J. Pharmacol. Exp. Ther. 279: 1422 (1996), Damaj etal., J. Pharmacol. Exp. Ther. 291: 390 (1999); Chiari et al.,Anesthesiology 91: 1447 (1999), Lavand'homme and Eisenbach,Anesthesiology 91: 1455 (1999), Holladay et al., J. Med. Chem. 40(28):4169-94 (1997), Bannon et al., Science 279: 77 (1998), PCT WO 94/08992,PCT WO 96/31475, PCT WO 96/40682, and U.S. Pat. Nos. 5,583,140 toBencherif et al., 5,597,919 to Dull et al., 5,604,231 to Smith et al.and 5,852,041 to Cosford et al.

CNS Disorders

(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine, a pharmaceuticallyacceptable salt thereof, or a pharmaceutical compositions containingsuch are useful in the treatment or prevention of a variety of CNSdisorders, including neurodegenerative disorders, neuropsychiatricdisorders, neurologic disorders, and addictions. The compounds and theirpharmaceutical compositions can be used to treat or prevent cognitiveimpairments and dysfunctions, age-related and otherwise; attentionaldisorders and dementias, including those due to infectious agents ormetabolic disturbances; to provide neuroprotection; to treat convulsionsand multiple cerebral infarcts; to treat mood disorders, compulsions andaddictive behaviors; to provide analgesia; to control inflammation, suchas mediated by cytokines and nuclear factor kappa B; to treatinflammatory disorders; to provide pain relief; and to treat infections,as anti-infectious agents for treating bacterial, fungal, and viralinfections. Among the disorders, diseases and conditions that thecompounds and pharmaceutical compositions of the present invention canbe used to treat or prevent are: age-associated memory impairment(AAMI), mild cognitive impairment (MCI), age-related cognitive decline(ARCD), pre-senile dementia, early onset Alzheimer's disease, seniledementia, dementia of the Alzheimer's type, Alzheimer's disease,cognitive impairment no dementia (CIND), Lewy body dementia,HIV-dementia, AIDS dementia complex, vascular dementia, Down syndrome,head trauma, traumatic brain injury (TBI), dementia pugilistica,Creutzfeld-Jacob Disease and prion diseases, stroke, ischemia, attentiondeficit disorder, attention deficit hyperactivity disorder, dyslexia,schizophrenia, schizophreniform disorder, schizoaffective disorder,cognitive dysfunction in schizophrenia, cognitive deficits inschizophrenia, Parkinsonism including Parkinson's disease,postencephalitic parkinsonism, parkinsonism-dementia of Gaum,frontotemporal dementia Parkinson's Type (FTDP), Pick's disease,Niemann-Pick's Disease, Huntington's Disease, Huntington's chorea,tardive dyskinesia, hyperkinesia, progressive supranuclear palsy,progressive supranuclear paresis, restless leg syndrome,Creutzfeld-Jakob disease, multiple sclerosis, amyotrophic lateralsclerosis (ALS), motor neuron diseases (MND), multiple system atrophy(MSA), corticobasal degeneration, Guillain-Barré Syndrome (GBS), andchronic inflammatory demyelinating polyneuropathy (CIDP), epilepsy,autosomal dominant nocturnal frontal lobe epilepsy, mania, anxiety,depression, premenstrual dysphoria, panic disorders, bulimia, anorexia,narcolepsy, excessive daytime sleepiness, bipolar disorders, generalizedanxiety disorder, obsessive compulsive disorder, rage outbursts,oppositional defiant disorder, Tourette's syndrome, autism, drug andalcohol addiction, tobacco addiction, and eating disorders.

Cognitive impairments or dysfunctions may be associated with psychiatricdisorders or conditions, such as schizophrenia and other psychoticdisorders, including but not limited to psychotic disorder,schizophreniform disorder, schizoaffective disorder, delusionaldisorder, brief psychotic disorder, shared psychotic disorder, andpsychotic disorders due to a general medical conditions, dementias andother cognitive disorders, including but not limited to mild cognitiveimpairment, pre-senile dementia, Alzheimer's disease, senile dementia,dementia of the Alzheimer's type, age-related memory impairment, Lewybody dementia, vascular dementia, AIDS dementia complex, dyslexia,Parkinsonism including Parkinson's disease, cognitive impairment anddementia of Parkinson's Disease, cognitive impairment of multiplesclerosis, cognitive impairment caused by traumatic brain injury,dementias due to other general medical conditions, anxiety disorders,including but not limited to panic disorder without agoraphobia, panicdisorder with agoraphobia, agoraphobia without history of panicdisorder, specific phobia, social phobia, obsessive-compulsive disorder,post-traumatic stress disorder, acute stress disorder, generalizedanxiety disorder and generalized anxiety disorder due to a generalmedical condition, mood disorders, including but not limited to majordepressive disorder, dysthymic disorder, bipolar depression, bipolarmania, bipolar I disorder, depression associated with manic, depressiveor mixed episodes, bipolar II disorder, cyclothymic disorder, and mooddisorders due to general medical conditions, sleep disorders, includingbut not limited to dyssomnia disorders, primary insomnia, primaryhypersomnia, narcolepsy, parasomnia disorders, nightmare disorder, sleepterror disorder and sleepwalking disorder, mental retardation, learningdisorders, motor skills disorders, communication disorders, pervasivedevelopmental disorders, attention-deficit and disruptive behaviordisorders, attention deficit disorder, attention deficit hyperactivitydisorder, feeding and eating disorders of infancy, childhood, or adults,tic disorders, elimination disorders, substance-related disorders,including but not limited to substance dependence, substance abuse,substance intoxication, substance withdrawal, alcohol-related disorders,amphetamine or amphetamine-like-related disorders, caffeine-relateddisorders, cannabis-related disorders, cocaine-related disorders,hallucinogen-related disorders, inhalant-related disorders,nicotine-related disorders, opioid-related disorders, phencyclidine orphencyclidine-like-related disorders, and sedative-, hypnotic- oranxiolytic-related disorders, personality disorders, including but notlimited to obsessive-compulsive personality disorder and impulse-controldisorders.

The above conditions and disorders are discussed in further detail, forexample, in the American Psychiatric Association: Diagnostic andStatistical Manual of Mental Disorders, Fourth Edition, Text Revision,Washington, DC, American Psychiatric Association, 2000.

Inflammation

The nervous system, primarily through the vagus nerve, is known toregulate the magnitude of the innate immune response by inhibiting therelease of macrophage tumor necrosis factor (TNF). This physiologicalmechanism is known as the “cholinergic anti-inflammatory pathway” (see,for example, Tracey, “The inflammatory reflex,” Nature 420: 853-9(2002)). Excessive inflammation and tumor necrosis factor synthesiscause morbidity and even mortality in a variety of diseases. Thesediseases include, but are not limited to, endotoxemia, rheumatoidarthritis, osteoarthritis, psoriasis, asthma, atherosclerosis,idiopathic pulmonary fibrosis, and inflammatory bowel disease.

Inflammatory conditions that can be treated or prevented byadministering the compounds described herein include, but are notlimited to, chronic and acute inflammation, psoriasis, endotoxemia,gout, acute pseudogout, acute gouty arthritis, arthritis, rheumatoidarthritis, osteoarthritis, allograft rejection, chronic transplantrejection, asthma, atherosclerosis, mononuclear-phagocyte dependent lunginjury, idiopathic pulmonary fibrosis, atopic dermatitis, chronicobstructive pulmonary disease, adult respiratory distress syndrome,acute chest syndrome in sickle cell disease, inflammatory bowel disease,irritable bowel syndrome, Crohn's disease, ulcerative colitis, acutecholangitis, aphteous stomatitis, pouchitis, glomerulonephritis, lupusnephritis, thrombosis, and graft vs. host reaction.

Inflammatory Response Associated with Bacterial and/or Viral Infection

Many bacterial and/or viral infections are associated with side effectsbrought on by the formation of toxins, and the body's natural responseto the bacteria or virus and/or the toxins. As discussed above, thebody's response to infection often involves generating a significantamount of TNF and/or other cytokines. The over-expression of thesecytokines can result in significant injury, such as septic shock (whenthe bacteria is sepsis), endotoxic shock, urosepsis viral pneumonitis,and toxic shock syndrome.

Cytokine expression is mediated by NNRs, and can be inhibited byadministering agonists or partial agonists of these receptors. Thosecompounds described herein that are agonists or partial agonists ofthese receptors can therefore be used to minimize the inflammatoryresponse associated with bacterial infection, as well as viral andfungal infections. Examples of such bacterial infections includeanthrax, botulism, and sepsis. Some of these compounds may also haveantimicrobial properties.

These compounds can also be used as adjunct therapy in combination withexisting therapies to manage bacterial, viral and fungal infections,such as antibiotics, antivirals and antifungals. Antitoxins can also beused to bind to toxins produced by the infectious agents and allow thebound toxins to pass through the body without generating an inflammatoryresponse. Examples of antitoxins are disclosed, for example, in U.S.Pat. No. 6,310,043 to Bundle et al. Other agents effective againstbacterial and other toxins can be effective and their therapeutic effectcan be complemented by co-administration with the compounds describedherein.

Pain

The compounds of the present invention and their pharmaceuticalcompositions are particularly useful in treating and preventing pain,including acute, persistent, and chronic pain. The pain types andpainful conditions that can be treated or prevented using the compoundsand their pharmaceutical compositions include nociceptive pain,neurologic pain, neuropathic pain, female-specific pain, inflammatorypain, fibromyalgia, post-operative pain, pain due to medical condition(such as AIDS or other disorder), arthritis pain, temporomandibularjoint disorder, burn pain, injury pain, back pain, sciatica, foot pain,headache, abdominal pain, muscle and connective tissue pain, joint pain,breakthrough pain, cancer pain, somatic pain, visceral pain, chronicfatigue syndrome, psychogenic pain, and pain disorder.

Neuropathic pain syndromes are the consequence of abnormal changesoccurring within pain signaling systems of both the peripheral andcentral nervous system. Their diverse etiology and symptomatology havetraditionally rendered them particularly difficult to treat with anyconsistency. Examples of neuropathic pain syndromes include thoseattributed to trigeminal or herpetic neuralgia, peripheral neuropathies(diabetic neuropathy, chemotherapy-induced neuropathy), post-herpeticneuralgia, entrapment neuropathies (carpel-tunnel syndrome),radiculopathy, complex regional pain syndrome, causalgia, low back pain,spontaneous pain (pain without an external stimulus), anddeafferentation syndromes such as brachial plexus avulsion and spinalcord injury. Hyperalgesia (strong pain associated with a mild stimulus),allodynia (pain due associated with an innocuous stimulus), parethesias(sensation of numbness or pricking in the absence of an externalstimulus), and dysesthesia (spontaneous or evoked unpleasant abnormalsensations) are also typically characterized as types of neuropathicpain. The compounds of the present invention and their pharmaceuticalcompositions are particularly useful in treating and preventing theseneuropathic pain types and associated conditions.

Other Disorders

In addition to treating CNS disorders, inflammation, and pain, thecompounds of the present invention can be also used to prevent or treatcertain other conditions, diseases, and disorders in which NNRs play arole. Examples include autoimmune disorders such as lupus, disordersassociated with cytokine release, cachexia secondary to infection (e.g.,as occurs in AIDS, AIDS related complex and neoplasia), obesity,pemphitis, urinary incontinence, retinal diseases, infectious diseases,myasthenia, Eaton-Lambert syndrome, hypertension, osteoporosis,vasoconstriction, vasodilatation, cardiac arrhythmias, type I diabetes,type II diabetes, bulimia, anorexia, diarrhea, constipation, and ulcers,as well as those indications set forth in published PCT application WO98/25619. The compounds of this invention can also be administered totreat convulsions such as those that are symptomatic of epilepsy, and totreat conditions such as syphillis and Creutzfeld-Jakob disease.

Diagnostic Uses

The compounds can be used in diagnostic compositions, such as probes,particularly when they are modified to include appropriate labels. Theprobes can be used, for example, to determine the relative number and/orfunction of specific receptors, particularly the α4β2 receptor subtype.For this purpose the compounds of the present invention most preferablyare labeled with a radioactive isotopic moiety such as ¹¹C, ¹⁸F, ⁷⁶Br,¹²³I or ¹²⁵I.

The administered compounds can be detected using known detection methodsappropriate for the label used. Examples of detection methods includeposition emission topography (PET) and single-photon emission computedtomography (SPECT). The radiolabels described above are useful in PET(e.g., 11C, ¹⁸F or ⁷⁶Br) and SPECT (e.g., ¹²³I) imaging, with half-livesof about 20.4 minutes for ¹¹C, about 109 minutes for ¹⁸F, about 13 hoursfor ¹²³I, and about 16 hours for ⁷⁶Br. A high specific activity isdesired to visualize the selected receptor subtypes at non-saturatingconcentrations. The administered doses typically are below the toxicrange and provide high contrast images. The compounds are expected to becapable of administration in non-toxic levels. Determination of dose iscarried out in a manner known to one skilled in the art of radiolabelimaging. See, for example, U.S. Pat. No. 5,969,144 to London et al.

The compounds can be administered using known techniques. See, forexample, U.S. Pat. No. 5,969,144 to London et al. The compounds can beadministered in formulation compositions that incorporate otheringredients, such as those types of ingredients that are useful informulating a diagnostic composition. Compounds useful in accordancewith carrying out the present invention most preferably are employed informs of high purity. See, U.S. Pat. No. 5,853,696 to Elmalch et al.

After the compounds are administered to a subject (e.g., a humansubject), the presence of that compound within the subject can be imagedand quantified by appropriate techniques in order to indicate thepresence, quantity, and functionality of selected NNR subtypes. Inaddition to humans, the compounds can also be administered to animals,such as mice, rats, dogs, and monkeys. SPECT and PET imaging can becarried out using any appropriate technique and apparatus. SeeVillemagne et al., In: Arneric et al. (Eds.) Neuronal NicotinicReceptors: Pharmacology and Therapeutic Opportunities, 235-250 (1998)and U.S. Pat. No. 5,853,696 to Elmalch et al.

The radiolabeled compounds bind with high affinity to selective NNRsubtypes (e.g., α4β2) and preferably exhibit negligible non-specificbinding to other nicotinic cholinergic receptor subtypes (e.g., thosereceptor subtypes associated with muscle and ganglia). As such, thecompounds can be used as agents for noninvasive imaging of nicotiniccholinergic receptor subtypes within the body of a subject, particularlywithin the brain for diagnosis associated with a variety of CNS diseasesand disorders.

In one aspect, the diagnostic compositions can be used in a method todiagnose disease in a subject, such as a human patient. The methodinvolves administering to that patient a detectably labeled compound asdescribed herein, and detecting the binding of that compound to selectedNNR subtypes (e.g., α4β2 receptor subtypes). Those skilled in the art ofusing diagnostic tools, such as PET and SPECT, can use the radiolabeledcompounds described herein to diagnose a wide variety of conditions anddisorders, including conditions and disorders associated withdysfunction of the central and autonomic nervous systems. Such disordersinclude a wide variety of CNS diseases and disorders, includingAlzheimer's disease, Parkinson's disease, and schizophrenia. These andother representative diseases and disorders that can be evaluatedinclude those that are set forth in U.S. Pat. No. 5,952,339 to Bencherifet al.

In another aspect, the diagnostic compositions can be used in a methodto monitor selective nicotinic receptor subtypes of a subject, such as ahuman patient. The method involves administering a detectably labeledcompound as described herein to that patient and detecting the bindingof that compound to selected nicotinic receptor subtypes namely, theα4β2 receptor subtypes.

Receptor Binding

The compounds of this invention can be used as reference ligands inbinding assays for compounds which bind to NNR subtypes, particularlythe α4β2 receptor subtypes. For this purpose the compounds of thisinvention are preferably labeled with a radioactive isotopic moiety suchas ³H, or ¹⁴C. Examples of such binding assays are described in detailbelow.

EXAMPLES

The following examples are provided to illustrate the present invention,and should not be construed as limiting thereof. In these examples, allparts and percentages are by weight, unless otherwise noted.

Example 1 Instrumentation and Experimental Protocols forCharacterization of Salt Forms of(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine

X-Ray Powder Diffraction (XRPD)

X-Ray Powder Diffraction patterns were collected on a Bruker AXS C2GADDS diffractometer using CuKa radiation (40 kV, 40 mA), automated XYZstage, laser video microscope for auto-sample positioning and a HiStar2-dimensional area detector. X-ray optics consists of a single Gobelmultilayer mirror coupled with a pinhole collimator of 0.3 mm. The beamdivergence (i.e. the effective size of the X-ray beam on the sample) wasapproximately 4 mm. A θ-θ continuous scan mode was employed with asample—detector distance of 20 cm which gives an effective 20 range of3.2°-29.7°. Typically the sample would be exposed to the X-ray beam for120 seconds. Samples run under ambient conditions were prepared as flatplate specimens using powder as received without grinding. Approximately1-2 mg of the sample was lightly pressed on a glass slide to obtain aflat surface. Samples run under non-ambient conditions were mounted on asilicon wafer with heat-conducting compound. The sample was then heatedto the appropriate temperature at ca. 10° C./min and subsequently heldisothermally for about 5 min before data collection was initiated. Peakpositions are reported as °2θ with an accuracy of ±0.1°.

Single Crystal XRD (SXD)

Data were collected on a Bruker AXS 1K SMART CCD diffractometer equippedwith an Oxford Cryosystems Cryostream cooling device. Structures weresolved using either the SHELXS or SHELXD programs and refined with theSHELXL program as part of the Bruker AXS SHELXTL suite. Unless otherwisestated, hydrogen atoms attached to carbon were placed geometrically andallowed to refine with a riding isotropic displacement parameter.Hydrogen atoms attached to a heteroatom were located in a differenceFourier synthesis and were allowed to refine freely with an isotropicdisplacement parameter.

Nuclear Magnetic Resonance (NMR) Spectrometry

NMR spectra were collected on either a Varian Unity 300 MHz instrumentor a Bruker 400 MHz instrument equipped with an auto-sampler andcontrolled by a DRX400 console. Automated experiments were acquiredusing ICONNMR v4.0.4 (build 1) running with Topspin v 1.3 (patch level8) using the standard Bruker loaded experiments. For non-routinespectroscopy, data were acquired through the use of Topspin alone.

Melting Point

A Fisher-Johns hot stage melting point apparatus was used, at a settingcorresponding to a heating rate of about 5° C. per min.

Differential Scanning Calorimetry (DSC)

DSC data were collected on a TA Instruments Q1000 or a Mettler DSC 823eequipped with a 50 position auto-sampler. The instrument was calibratedfor energy and temperature calibration using certified indium. Typically0.5-1.5 mg of each sample, in a pin-holed aluminium pan, was heated at10° C./min from 25° C. to 175-200° C. A nitrogen purge at 30 mL/min wasmaintained over the sample.

Thermo-Gravimetric Analysis (TGA)

TGA data were collected on a TA Instruments Q500 TGA equipped with a 16position auto-sampler or a Mettler TGA/SDTA 851 e equipped with a 34position auto sampler. TA Instruments Q500: The instrument wastemperature calibrated using certified Alumel. Typically 5-10 mg of eachsample was loaded onto a pre-tared platinum crucible and aluminium DSCpan, and was heated at 10° C./min from ambient temperature to 350° C. Anitrogen purge at 60 mL/min was maintained over the sample. MettlerTGA/SDTA 851e: The instrument was temperature calibrated using certifiedindium. Typically 5-10 mg of each sample was loaded onto a pre-taredaluminum crucible and was heated at 10° C./min from ambient temperatureto 350° C. A nitrogen purge at 50 mL/min was maintained over the sample.

Polarized Light Microscopy (PLM)

Samples were studied on a Leica LM/DM polarized light microscope with adigital video camera for image capture. A small amount of each samplewas placed on a glass slide, mounted in immersion oil and covered with aglass slip, the individual particles being separated as well aspossible. The sample was viewed with appropriate magnification andpartially polarised light, coupled to a A false-color filter.

Hot Stage Microscopy (HSM)

Hot Stage Microscopy was carried out using a Leica LM/DM polarized lightmicroscope combined with a Mettler-Toledo MTFP82HT hot-stage and adigital video camera for image capture. A small amount of each samplewas placed onto a glass slide with individual particles separated aswell as possible. The sample was viewed with appropriate magnificationand partially polarized light, coupled to a A false-color filter, whilstbeing heated from ambient temperature typically at 10° C./min.

Dynamic Vapor Sorption (DVS)

Sorption isotherms were determined using a SMS DVS Intrinsic moisturesorption analyzer controlled by SMS Analysis suite software. The sampletemperature was maintained at 25° C. by the instrument controls. Thehumidity was controlled by mixing streams of dry and wet nitrogen, witha total flow rate of 200 mL/min. The relative humidity was measured by acalibrated Rotronic probe (dynamic range of 1.0-100% RH), located nearthe sample. The weight change, (mass relaxation) of the sample as afunction of % RH was constantly monitored by the microbalance (accuracy±0.005 mg).

Typically a 5-20 mg sample was placed on the tared mesh stainless steelbasket under ambient conditions. The sample was loaded and unloaded at40% RH and 25° C. (typical ambient conditions). A moisture sorptionisotherm was performed as outlined below (2 scans giving 1 completecycle). The standard isotherm was performed at 25° C. at 10% RHintervals over a 0-90% RH range.

DVS Generic Method Parameters

Parameters Values Adsorption - Scan 1 40-90 Desorption/Adsorption - Scan2 90-Dry, Dry-40 Intervals (% RH) 10 Number of Scans 2 Flow rate(mL/min) 200 Temperature (° C.) 25 Stability (° C./min) 0.2 SorptionTime (hours) 6 hour time out

Samples were recovered after completion of the isotherm and re-analyzedby XRPD.

Water Determination by Karl Fischer (KF)

The water content of each sample was measured on a Mettler Toledo DL39Coulometer using Hydranal Coulomat AG reagent and an argon purge.Weighed solid samples were introduced into the vessel on a platinum TGApan which was connected to a subaseal to avoid water ingress. Approx 10mg of sample was used per titration and duplicate determinations weremade.

Thermodynamic Aqueous Solubility by HPLC

Aqueous solubility was determined by suspending sufficient compound inwater to give a maximum final concentration of 10 mg/mL of the parentfree-form of the compound. The suspension was equilibrated at 25° C. for24 h, and then the pH was measured. The suspension was then filteredthrough a glass fiber C filter into a 96 well plate. The filtrate wasthen diluted by a factor of 101. Quantitation was by HPLC with referenceto a standard solution of approximately 0.1 mg/mL in DMSO. Differentvolumes of the standard, diluted and undiluted sample solutions wereinjected. The solubility was calculated using the peak areas determinedby integration of the peak found at the same retention time as theprincipal peak in the standard injection. If there was sufficient solidin the filter plate, the XRPD was collected.

HPLC method parameters for thermodynamic aqueous solubility method

Type of method: Reverse phase with gradient elution Column: PhenomenexLuna, C18 (2) 5 μm, 50 × 4.6 mm Column Temperature (° C.): 25 StandardInjections (μL): 1, 2, 3, 5, 7, 10 Test Injections (μL): 1, 2, 3, 10,20, 50 Detection: 260, 80 Wavelength, Bandwidth (nm): Flow Rate(mL/min): 2 Phase A:  0.1% TFA in water Phase B: 0.085% TFA inacetonitrile Time (min) % Phase A % Phase B Timetable: 0.0 95 5 1.0 8020 2.3 5 95 3.3 5 95 3.5 95 5 4.4 95 5

Analysis was performed on an Agilent HP1100 series system equipped witha diode array detector and using ChemStation software vB.02.01-SR1.

Chemical Purity by HPLC

Purity analysis was performed on an Agilent HP1100 series systemequipped with a diode array detector and using ChemStation softwarevB.02.01-SR1.

HPLC method parameters for chemical purity determination

Sample Preparation 0.5 mg/mL in acetonitrile:water 1:1 (v/v) Column:Phenomenex Luna C18 (2), 150 × 4.6 mm, 5 μm Column Temperature (° C.):25 Injection (μL):  5 Detection: 255, 90 Wavelength, Bandwidth(nm): FlowRate (mL/min):  1 Phase A:  0.1% TFA in water Phase B: 0.085% TFA inacetonitrile Time (min) % Phase A % Phase B Timetable: 0 95 5 25 5 9525.2 95 5 30 95 5Ion Chromatography

Data were collected on a Metrohm 761 Advanced Compact IC (for cations)and a Metrohm 861 Advanced Compact IC (for anions) using IC Net softwarev2.3. Samples were prepared as 1000 ppm stocks in DMSO. Samples werediluted to 100 ppm with DMSO prior to testing. Quantification wasachieved by comparison with standard solutions of known concentration ofthe ion being analyzed.

Ion Chromatography Method for Anions

Type of method Anion exchange Column: Metrosep A Supp 5-250 (4.0 × 250mm) Column Temperature (° C.): Ambient Injection (μL): 20 Detection:Conductivity detector Flow Rate (mL/min): 0.7 Eluent: 3.2 mM sodiumcarbonate, 1.0 mM sodium hydrogen carbonate in waterIon Chromatography Method for Cations

Type of method Cation exchange Column: Metrosep C 2-250 (4.0 × 250 mm)Column Temperature Ambient (° C.): Injection (μL): 20 Detection:Conductivity detector Flow Rate (mL/min): 1.0 Eluent:  4.0 mM Tartaricacid, 0.75 mM Dipicolinic acid in waterpKa Determination and Prediction

Data were collected on a Sirius GlpKa instrument with a D-PASattachment. Measurements were made at 25° C. in aqueous solution by UVand in methanol water mixtures by potentiometry. The titration media wasionic-strength adjusted (ISA) with 0.15 M KCl (aq). The values found inthe methanol water mixtures were corrected to 0% co-solvent viaYasuda-Shedlovsky extrapolation. The data were refined using RefinementPro software v1.0. Prediction of pKa values was made using ACD pKaprediction software v9.

Log P Determination

Data were collected by potentiometric titration on a Sirius GlpKainstrument using three ratios of octanol:ionic-strength adjusted (ISA)water to generate Log P, Log P_(ion), and Log D values. The data wererefined using Refinement Pro software v1.0. Prediction of Log P valueswas made using ACD v9 and Syracuse KOWWIN v1.67 software.

Example 2 Synthesis of Tert-Butyl(R)-3-(methylsulfonyloxy)pyrrolidine-1-carboxylate (2)

Procedure A: To a solution of tert-butyl(R)-3-hydroxypyrrolidine-1-carboxylate (200 g, 1.07 mol) andtriethylamine (167 g, 1.63 mol) in toluene (700 mL) at -20 to -30° C.was added methanesulfonyl chloride (156 g, 1.36 mol) drop-wise whilemaintaining the temperature at -10 to -20° C. The solution was warmed toambient temperature and allowed to stir. The reaction solution wassampled hourly and analyzed by HPLC to establish completion of thereaction. Upon completion of the reaction, the suspension was filteredto remove the triethylamine hydrochloride. The filtrate was washed with˜600 mL of dilute aqueous sodium bicarbonate solution. The organic layerwas dried and concentrated under reduced pressure to give 2 as a viscousoil (260 g, 92%) which is used without further purification. ¹H NMR(CDCl₃, 400 MHz) δ 5.27 (m, 1 H), 3.44-3.76 (m, 4H), 3.05 (s, 3H), 2.26(m, 1H), 2.15 (m, 1H), 1.47 (s, 9H).

Procedure B: A reactor was charged with tert-butyl(R)-3-hydroxypyrrolidine-1-carboxylate (2.00 kg, 10.7 mol), toluene(8.70 kg) and triethylamine (1.75 kg, 17.3 mol). The reactor was flushedwith nitrogen for 15 min. The mixture was stirred and cooled to 3° C.Methanesulfonyl chloride (1.72 kg, mol) was slowly added (over a 2 hperiod) with continuous ice bath cooling (exothermic reaction) (aftercomplete addition, the temperature was 14° C.). The mixture, now viscouswith precipitated triethylamine hydrochloride, was stirred 12 h as itwarmed to 20° C. Both GC and TLC analysis (ninhydrin stain) indicatedthat no starting material remained. The mixture was filtered to removethe triethylamine hydrochloride, and the filtrate was returned to thereactor. The filtrate was then washed (2×3 kg) with 5% aqueous sodiumbicarbonate, using 15 min of stirring and 15 min of settling time foreach wash. The resulting organic layer was dried over anhydrous sodiumsulfate and filtered. The volatiles were removed from the filtrate undervacuum, first at 50° C. for 4 h and then at ambient temperature for 10h. The residue weighed 3.00 kg (106% yield) and was identical bychromatographic and

NMR analysis to previously prepared samples, with the exception that itcontained toluene.

Example 3 Synthesis of diethyl(R)-2-(1-(tert-butoxycarbonyl)pyrrolidin-3-yl)malonate (3)

Preparation A: To a solution of potassium tert-butoxide (187 g, 1.62mol) in 1-methyl-2-pyrrolidinone (1.19 L) was added diethyl malonate(268 g. 1.67 mol) while maintaining the temperature below 35° C. Thesolution was heated to 40° C. and stirred for 20-30 min. tert-Butyl(R)-3-(methylsulfonyloxyl)pyrrolidine-1-carboxylate (112 g, 420 mmol)was added and the solution was heated to 65° C. and stirred for 6 h .The reaction solution was sampled every 2 h and analyzed by HPLC toestablish completion of the reaction. Upon completion of reaction (10-12h), the mixture was cooled to around 25° C. De-ionized water (250 mL)was added to the solution, and the pH was adjusted to 3-4 by addition of2N hydrochloric acid (650 mL). The resulting suspension was filtered,and water (1.2 L) and chloroform (1.4 L) were added. The solution wasmixed thoroughly, and the chloroform layer was collected and evaporatedunder reduced pressure to give a yellow oil. The oil was dissolved inhexanes (2.00 L) and washed with deionized water (2×1.00 L). The organiclayer was concentrated under reduced pressure at 50-55° C. to give apale yellow oil (252 g) which ¹H NMR analysis indicates to be 49.1% of 3(123.8 g) along with 48.5% diethyl malonate (122g), and 2% of1-methyl-2-pyrrolidinone (5 g). The material was carried forward to thenext step without further purification. ¹H NMR (CDCl₃, 400 MHz) δ 4.20(q, 4H), 3.63 (m, 1H), 3.48 (m, 1H), 3.30 (m, 1H), 3.27 (d, J =10 Hz,1H), 3.03 (m, 1H), 2.80 (m, 1H), 2.08 (m, 1H), 1.61 (m,1H), 1.45 (s,9H), 1.27 (t, 6H).

Preparation B: A reactor, maintained under a nitrogen atmosphere, wascharged with 200 proof ethanol (5.50 kg) and 21% (by weight) sodiumethoxide in ethanol (7.00 kg, 21.6 mol). The mixture was stirred andwarmed to 30° C. Diethyl malonate (3.50 kg, 21.9 mol) was added over a20 min period. The reaction mixture was then warmed at 40° C. for 1.5 h.A solution of tert-butyl(R)-3-(methylsulfonyloxyl)pyrrolidine-1-carboxylate (3.00 kg of theproduct from Example 2, Procedure B, 10.7 mol) in 200 proof ethanol(5.50 kg) was added, and the resulting mixture was heated at reflux (78°C.) for 2 h. Both GC and TLC analysis (ninhydrin stain) indicated thatno starting material remained. The stirred mixture was then cooled to25° C., diluted with water (2.25 kg), and treated slowly with a solutionof concentrated hydrochloric acid (1.27 kg, 12.9 mol) in water (5.44kg). This mixture was washed twice with methyl tert-butyl ether (MTBE)(14.1 kg and 11.4 kg), using 15 min of stirring and 15 min of settlingtime for each wash. The combined MTBE washes were dried over anhydroussodium sulfate (1 kg), filtered and concentrated under vacuum at 50° C.for 6 h. The residue (red oil) weighed 4.45 kg and was 49% desiredproduct by GC analysis (62% overall yield from tert-butyl(R)-3-hydroxypyrrolidine-1-carboxylate).

Example 4 Synthesis of(R)-2-(1-(tert-butoxycarbonyl)pyrrolidin-3-yl)malonic acid (4)

Procedure A: To a solution of the product of Example 3, Procedure A (232g), containing 123.8 g (380 mmol) of 3 and 121.8 g (760 mmol) of diethylmalonate, in tetrahydrofuran (1.2 L) was added a 21% potassium hydroxidesolution (450 g in 0.50 L of deionized water) while maintaining thetemperature below 25° C. The reaction mixture was heated to 45° C. andstirred for 1 h. The reaction solution was sampled every hour andanalyzed by HPLC to establish completion of the reaction. Uponcompletion of reaction (2-3 h), the mixture was cooled to around 25° C.The aqueous layer was collected and cooled to 5° C. The pH was adjustedto 2 by addition of 4N hydrochloric acid (750 mL), and the resultingsuspension was held at 5-10° C. for 30 min. The mixture was filtered,and the filter cake was washed with hexanes (1 L). The aqueous filtratewas extracted with chloroform (1 L) and the chloroform layer was putaside. The solids collected in the filtration step were re-dissolved inchloroform (1 L) by heating to 40° C. The solution was filtered toremove un-dissolved inorganic solids. The chloroform layers werecombined and concentrated under reduced pressure at 50-55° C. to give anoff-white solid (15 g). The solids were combined and dissolved in ethylacetate (350 mL) to give a suspension that was warmed to 55-60° C. for 2h. The suspension was filtered while hot and the resulting cake washedwith ethyl acetate (2×150 mL) and hexanes (2×250 mL) to give 83.0 g(80.1%) of 4 as a white solid which was used in the next step withoutfurther purification. ¹H NMR (d₄-CH₃OH, 400 MHz) δ 3.60 (m, 1H), 3.46(m, 1H), 3.29-3.32 (m, 2H), 2.72 (m, 1H), 2.09 (m, 1H), 1.70 (m, 1H),1.45 (s, 9H).

Procedure B: A solution of the product of Example 3, Procedure B (4.35kg), containing 2.13 kg (6.47 mol) of 3, in tetrahydrofuran (13.9 kg)was added to a stirred, cooled solution of potassium hydroxide (1.60 kg,40.0 mol) in deionized water (2.00 kg) under a nitrogen atmosphere,while maintaining the temperature below 35° C. The reaction mixture washeated and maintained at 40-45° C. for 24 h, by which time GC and TLCanalysis indicated that the reaction was complete. The mixture wascooled to 25° C. and washed with MTBE (34 kg), using 15 min of stirringand 15 min of settling time. The aqueous layer was collected and cooledto 1° C. A mixture of concentrated hydrochloric acid (2.61 kg, 26.5 mol)in deionized water (2.18 kg) was then added slowly, keeping thetemperature of the mixture at <15° C. during and for 15 min after theaddition. The pH of the solution was adjusted to 3.7 by further additionof hydrochloric acid. The white solid was collected by filtration,washed with water (16 kg), and vacuum dried at ambient temperature for 6d. The dry solid weighed 1.04 kg. The filtrate was cooled to <10° C. andkept at that temperature as the pH was lowered by addition of morehydrochloric acid (1.6 L of 6 N was used; 9.6 mol; final pH=2). Thewhite solid was collected by filtration, washed with water (8 L), andvacuum dried at 40° C. for 3 d. The dry solid weighed 0.25 kg. Thecombined solids (1.29 kg, 73% yield) were chromatographically identicalto previously prepared samples.

Example 5 Synthesis of(R)-2-(1-(tert-butoxycarbonyl)pyrrolidine-3-yl)acetic acid (5)

Procedure A: A solution of(R)-2-(1-(tert-butoxycarbonyl)pyrrolidin-3-yl)malonic acid (83 g) in1-methyl-2-pyrrolidinone (0.42 L) was stirred under nitrogen at 110-112°C. for 2 h . The reaction solution was sampled every hour and analyzedby HPLC to establish completion of the reaction. Upon completion ofreaction the reaction solution was cooled to 20-25° C. The solution wasmixed with de-ionized water (1.00 L), and

MTBE (1.00 L) was added. The phases were separated, and the organiclayer was collected. The aqueous phase was extracted with MTBE (1.00 L),then chloroform (1.00 L). The organic layers were combined andconcentrated under reduced pressure at 50-55° C. to give an oil. Thisoil was dissolved in MTBE (2.00 L) and washed twice with 0.6Nhydrochloric acid (2×1.00 L). The organic layer was collected andconcentrated under reduced pressure at 50-55° C. to give a semi-solid.The semi-solid was suspended in 1:4 ethyl acetate/hexanes (100 mL),heated to 50° C., held for 30 min, cooled to −10° C., and filtered. Thefiltrate was concentrated under reduced pressure to give an oil, whichwas dissolved in MTBE (250 mL) and washed twice with 0.6N hydrochloricacid (2×100 mL). The organic layer was concentrated under reducedpressure at 50-55° C. to give a semi-solid which was suspended in 1:4ethyl acetate/hexanes (50 mL), heated to 50° C., held for 30 min, cooledto −10° C., and filtered. The solids were collected, suspended inhexanes (200 mL), and collected by filtration to give 54.0 g (77.6%) of5. ¹H NMR (CDCl₃, 400 MHz) δ 11.00 (br s, 1 H), 3.63 (m, 1H), 3.45 (M,1H), 3.30 (M, 1H), 2.97 (m, 1H), 2.58 (m, 1H), 2.44 (m, 2H), 2.09 (m,1H), 1.59 (M, 1H), 1.46 (s, 9H).

Procedure B: A solution of(R)-2-(1-(tert-butoxycarbonyl)pyrrolidin-3-yl)malonic acid (1.04 kg,3.81 mol) in 1-methyl-2-pyrrolidinone (6.49 kg) was stirred undernitrogen at 110° C. for 5 h , by which time TLC and HPLC analysisindicated that the reaction was complete. The reaction mixture wascooled to 25° C. (4 h) and combined with water (12.8 kg) and MTBE (9.44kg). The mixture was stirred vigorously for 20 min, and the phases wereallowed to separate (10 h). The organic phase was collected, and theaqueous phase was combined with MTBE (9.44 kg), stirred for 15 min, andallowed to settle (45 min). The organic phase was collected, and theaqueous phase was combined with MTBE (9.44 kg), stirred for 15 min, andallowed to settle (15 min). The three organic phases were combined andwashed three times with 1N hydrochloric acid (8.44 kg portions) and oncewith water (6.39 kg), using 15 min of stirring and 15 min of settlingtime for each wash. The resulting solution was dried over anhydroussodium sulfate (2.0 kg) and filtered. The filtrate was concentratedunder reduced pressure at 31° C. (2 h) to give an solid. This solid washeated under vacuum for 4 h at 39° C. for 4 h and for 16 h at 25° C.,leaving 704 g (81%) of 5 (99.7% purity by GC).

Procedure C (streamlined synthesis of 5, using 2 as starting material):A stirred mixture of sodium ethoxide in ethanol (21 weight percent, 343g, 1.05 mol), ethanol (anhydrous, 300 mL) and diethyl malonate (168 g,1.05 mol) was heated to 40° C. for 1.5 h. To this mixture was added asolution of (R)-tert-butyl3-(methylsulfonyloxy)pyrrolidine-1-carboxylate (138 g, 0.592 mol) inethanol (100 mL) and the reaction mixture was heated to 78° C. for 8 h.The cooled reaction mixture was diluted with water (2.0 L) and acidifiedto pH =3 with 6M HCl (100 mL). The aqueous ethanol mixture was extractedwith toluene (1.0 L), and the organic phase concentrated under vacuum toafford 230 g of a red oil. The red oil was added at 85° C. to a 22.5weight percent aqueous potassium hydroxide (748 g, 3.01 mol). After theaddition was complete, the reaction temperature was allowed to slowlyrise to 102° C. while a distillation of ethanol ensued. When thereaction temperature had reached 102° C., and distillation had subsided,heating was continued for an additional 90 min. The reaction mixture wascooled to ambient temperature and washed with toluene (2×400 mL). To theaqueous layer was added 600 mL 6M hydrochloric acid, while keeping theinternal temperature below 20° C. This resulted in the formation of aprecipitate, starting at pH of about 4-5. The suspension was filtered,and the filter cake was washed with 300 mL water. The solid was driedunder vacuum to afford 77 g of(R)-2-(1-(tert-butoxycarbonyl)pyrrolidin-3-yl)malonic acid as anoff-white solid (54% yield with respect to (R)-tert-butyl3-(methylsulfonyloxy)pyrrolidine-1-carboxylate). ¹H NMR (DMSO-d₆, 400MHz): δ 3.47 (m, 1H); 3.32 (m, 1H); 3.24 (m, 1H); 3.16 (m, 1H); 3.92 (m,1H); 2.86 (m, 1H); 1.95 (m, 1H); 1.59 (m, 1H); 1.39 (s, 9H).

A suspension of (R)-2-(1-(tert-butoxycarbonyl)pyrrolidin-3-yl)malonicacid (15 g, 55 mmol) in toluene (150 mL) and dimethylsulfoxide (2 mL)was heated to reflux for a period of 2 h. The mixture was allowed toreach ambient and diluted with MTBE (150 mL). The organic solution waswashed with 10% aqueous citric acid (2×200 mL), and the solvent wasremoved under vacuum to afford 11.6 g of(R)-2-(1-(tert-butoxycarbonyl)-pyrrolidin-3-yl)acetic acid as anoff-white solid (92% yield). ¹H NMR (DMSO-d₆, 400 MHz): δ 12.1 (s, 1H);3.36-3.48 (m, 1H); 3.20-3.34 (m, 1H); 3.05-3.19 (m, 1H; 2.72-2.84 (m,1H); 2.30-2.42 (m, 1H), 2.22-2.30 (m, 2H); 1.85-2.00 (m, 1H); 1.38-1.54(m, 1H), 1.35 (2, 9H).

Example 6 Synthesis of tert-butyl(R)-3-(2-hydroxyethyl)pyrrolidine-1-carboxylate (6)

Procedure A: A solution of(R)-2-(1-(tert-butoxycarbonyl)pyrrolidine-3-yl)acetic acid (49.0 g, 214mmol) in tetrahydrofuran (THF) (200 mL) was cooled to −10° C. 250 mL(250 mmol) of a 1M borane in THF solution was added slowly to the flaskwhile maintaining the temperature lower than 0° C. The solution waswarmed to ambient temperature and stirred for 1 h. The solution wassampled hourly and analyzed by HPLC to establish completion of thereaction. Upon completion of the reaction, the solution was cooled to 0°C., and a 10% sodium hydroxide solution (80 mL) was added drop-wise overa 30 minute period to control gas evolution. The solution was extractedwith 500 mL of a 1:1 hexanes/ethyl acetate solution. The organic layerwas washed with saturated sodium chloride solution and dried with 10 gof silica gel. The silica gel was removed by filtration and washed with100 mL of 1:1 hexanes/ethyl acetate. The organic layers were combinedand concentrated under vacuum to give 6 (42 g, 91.3%) as a light-orangeoil that solidified upon sitting.¹H NMR (CDCl₃, 400 MHz) δ 3.67 (m, 2H),3.38-3.62 (m, 2H), 3.25 (m, 1H), 2.90 (m, 1H), 2.25 (m, 1H), 1.98-2.05(m, 1H), 1.61-1.69 (m, 2H), 1.48-1.59 (m, 2H), 1.46 (s, 9H).

Procedure B: Borane-THF complex (3.90 kg or L of 1M in THF, mol) wasadded slowly to a stirred solution of(R)-2-(1-(tert-butoxycarbonyl)pyrrolidine-3-yl)acetic acid (683 g, 3.03mol) in THF (2.5 kg), kept under nitrogen gas, and using a water bath tokeep the temperature between 23 and 28° C. The addition took 1.75 h.Stirring at 25° C. was continued for 1 h, after which time GC analysisindicated complete reaction. The reaction mixture was cooled to <10° C.and maintained below 25° C. as 10% aqueous sodium hydroxide (1.22 kg)was slowly added. The addition took 40 min. The mixture was stirred 1 hat 25° C., and then combined with 1:1 (v/v) heptane/ethyl acetate (7 L).

The mixture was stirred for 15 min and allowed to separate into phases(1 h). The organic phase was withdrawn, and the aqueous phase wascombined with a second 7 L portion of 1:1 heptane/ethyl acetate. Thiswas stirred for 15 min and allowed to separate into phases (20 min). Theorganic phase was again withdrawn, and the combined organic phases werewashed with saturate aqueous sodium chloride (4.16 kg), using 15 min ofmixing and 1 h of settling time. The organic phase was combined withsilica gel (140 g) and stirred 1 h. The anhydrous sodium sulfate (700 g)was added, and the mixture was stirred for 1.5 h. The mixture wasfiltered, and the filter cake was washed with 1:1 heptane/ethyl acetate(2 L). The filtrate was concentrated under vacuum at <40° C. for 6 h.The resulting oil weighed 670 g (103% yield) and contains traces ofheptane, but is otherwise identical to previously prepared samples of 6,by NMR analysis.

Example 7 Tert-butyl(R)-3-(2-(methylsulfonyloxy)ethyl)pyrrolidine-1-carboxylate (⁷)

Procedure A: To a solution of tert-butyl(R)-3-(2-hydroxymethyl)pyrrolidine-1-carboxylate (41.0 g, 190 mmol)) wasadded triethylamine (40 mL) in toluene (380 mL) and cooled to −10° C.Methanesulfonyl chloride (20.0 mL, 256 mmol) was added slowly so as tomaintain the temperature around −5 to 0° C. The solution was warmed toambient temperature and stirred for 1 h. The solution was sampled hourlyand analyzed by HPLC to establish completion of the reaction. Uponcompletion of reaction, the solution was filtered, and the filtrate waswashed with a 5% sodium bicarbonate solution (250 mL). The organic layerwas collected and washed with a saturated aqueous sodium chloridesolution (250 mL). The organic layer was collected, dried over silicagel (10 g), and concentrated under vacuum to give 7 (53.0 g, 92.8%) as alight-yellow viscous oil. ¹H NMR (CDCl₃, 400 MHz) δ 4.26 (t, J=6.8 Hz,2H), 3.41-3.63 (m, 2H), 3.27 (m, 1H), 3.02 (s, 3H), 2.92 (m, 1H), 2.28(m, 1H), 2.05 (m, 1H), 1.83 (m, 2H), 1.50-1.63 (m, 1H), 1.46 (s, 9H).

Procedure B: Under a nitrogen atmosphere, a solution of triethylamine(460 g, 4.55 mol) and tert-butyl(R)-3-(2-hydroxymethyl)pyrrolidine-1-carboxylate (the entire sample fromExample 7, Procedure B, 3.03 mol) in toluene (5.20 kg) was stirred andcooled to 5° C. Methanesulfonyl chloride (470 g, 4.10 mol) was addedslowly, over a 1.25 h, keeping the temperature below 15° C. using icebath cooling. The mixture was gradually warmed (over 1.5 h) to 35° C.,and this temperature was maintained for 1.25 h, at which point GCanalysis indicated that the reaction was complete. The mixture wascooled to 25° C., and solids were filtered off and the filter cakewashed with toluene (1.28 kg). The filtrate was stirred with 10% aqueoussodium bicarbonate (4.0 kg) for 15 min, and the phases were allowed toseparate for 30 min. The organic phase was then stirred with saturatedaqueous sodium chloride (3.9 kg) for 30 min, and the phases were allowedto separate for 20 min. The organic phase was combined with silica gel(160 g) and stirred for 1 h. Anhydrous sodium sulfate (540 g) was added,and the mixture was stirred an additional 40 min. The mixture was thenfiltered, and the filter cake was washed with toluene (460 g). Thefiltrate was concentrated under vacuum at 50° C. for 5 h, and theresulting oil was kept under vacuum at 23° C. for an additional 8h. Thisleft 798 g of 7, 93% pure by GC analysis.

Example 8 Synthesis of tert-butyl (R)-3-vinylpyrrolidine-1-carboxylate(9)

Procedure A: A solution of tert-butyl(R)-3-((methylsulfonyloxy)ethyl)pyrrolidine-1-carboxylate (49.0 g, 167mmol), sodium iodide (30.0 g, 200 mmol) and 1,2-dimethoxyethane (450 mL)was stirred at 50-60° C. for 4 h . The solution was sampled hourly andanalyzed by HPLC to establish completion of the reaction. Uponcompletion of reaction, the solution was cooled to −10° C., and solidpotassium tert-butoxide (32.0 g, 288 mmol) was added while maintainingtemperature below 0° C. The reaction mixture was warmed to ambienttemperature and stirred for 1 h. The mixture was sampled hourly andanalyzed by HPLC to establish completion of the reaction. Uponcompletion of reaction, the mixture was filtered through a pad ofdiatomaceous earth (25 g dry basis). The cake was washed with1,2-dimethoxyethane (100 mL). The combined filtrates were concentratedunder vacuum, to yield an orange oil with suspended solids. The oil wasdissolved in hexanes (400 mL), stirred for 30 min, and filtered toremove the solids. The organic layer was dried over silica gel (10 g),and concentrated under vacuum to give 9 (26.4 g, 82.9%) as a colorlessoil. ¹H NMR (CDCl₃, 400 MHz) δ 5.77 (m, 1H), 5.10 (dd, J=1.2 Hz, J=16Hz, 1H), 5.03 (dd, J=1.2 Hz, J=8.8 Hz, 1H), 3.41-3.59 (m, 2H), 3.29 (m,1H), 3.05 (m, 1H), 2.78 (m, 1H), 2.01 (m, 1H), 1.62-1.73 (m, 1H), 1.46(m, 9H).

Procedure B: A solution of tert-butyl(R)-3-(2-(methylsulfonyloxy)ethyl)pyrrolidine-1-carboxylate (792 g ofthe product of Example 7,

Procedure B, ˜2.5 mol), sodium iodide (484 g, 3.27 mol) and1,2-dimethoxyethane (7.2 L) was stirred at 55° C. for 4.5 h undernitrogen, at which time GC analysis indicated that the reaction wascomplete. The solution was cooled to <10° C., and solid potassiumtert-butoxide (484 g, 4.32 mol) was added in portions (1.25 h additiontime) while maintaining temperature below 15° C. The reaction mixturewas stirred 1 h at 5° C., warmed slowly (6 h) to 20° C., and stirred at20° C. for 1 h. The solution was filtered through a pad of diatomaceousearth (400 g dry basis). The filter cake was washed with1,2-dimethoxyethane (1.6 kg). The combined filtrates were concentratedunder vacuum, and the semisolid residue was stirred with heptane (6.0 L)for 2h. The solids were removed by filtration (the filter cake waswashed with 440 mL of heptane), and the filtrate was concentrated undervacuum at 20° C. to give 455 g of 9 (90.7% pure). A sample of thismaterial (350 g) was fractionally distilled at 20-23 torr to give 296 gof purified 9 (bp 130-133° C.) (>99% pure by GC analysis).

Example 9 Synthesis of (R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine (11)

Nitrogen was bubbled through a solution of (R)-tert-butyl3-vinylpyrrolidine-1-carboxylate (25 g, 127 mmol), 5-bromopyrimidine(30.3 g, 190 mmol), 1, 1′-bis(diphenylphosphino)ferrocene (2.11 g, 3.8mmol), and sodium acetate (18.8 gr, 229 mmol) in N,N-dimethylacetamide(250 mL) for 1 h, and palladium acetate (850 mg, 3.8 mmol) was added.The reaction mixture was heated to 150° C. at a rate of 40° C./h andstirred for 16 h. The mixture was cooled to 10° C. and quenched withwater (750 mL) while maintaining an internal temperature below 20° C.MTBE (300 mL) was added, followed by diatomaceous earth (40 g, drybasis). The suspension was stirred for 1 h at ambient temperature andfiltered through a bed of diatomaceous earth. The residue was washedwith MTBE (2×100 mL) and the filtrate was transferred to a 2-L vesselequipped with an overhead stirrer and charged with activated charcoal(40 g). The suspension was stirred for 2 h at ambient temperature andfiltered through diatomaceous earth. The residue was washed with MTBE(2×100 mL,), and the filtrate was concentrated in vacuo to afford 28.6 gof an orange oil. The oil is dissolved in MTBE (100 mL) and Si-Thiol ®(2.0 g, 1.46 mmol thiol/g, Silicycle Inc.) was added. The suspension wasstirred under nitrogen at ambient temperature for 3 h, filtered througha fine filter, and held in a glass container.

To a solution of 6 M HCl (70 mL) was added the filtrate over a period of30 min while maintaining the internal temperature between 20° C. and 23°C. The mixture was stirred vigorously for 1 h and the organic layerremoved. The remaining aqueous layer was basified with 45 wt % KOH (50mL), and the resulting suspension was extracted once with chloroform(300 mL). Evaporation of the solvent in vacuo (bath temperature at 45°C.) gave 16.0 g (71.8%), of (R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidinefree base as a red oil, which is immediately dissolved in isopropanol(50 mL) and used for salt formation.

Example 10 Synthesis of (R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidinemono-citrate

To a solution of citric acid (17.6 g, 91.6 mmol) in isopropanol (250 mL)and water (25 mL) was added drop-wise a solution of(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine free base (16.0 g, 91.2mmol) in isopropanol (50 mL) at 55° C. The resulting solution was seededwith (R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine mono-citrate Form II(200 mg) and stirred for 15 min. The suspension was heated to 65° C. andstirred for 1 h, after which the suspension was cooled to 20° C. at −10°C./h and allowed to stand at 20° C. for 12 h. The suspension wasfiltered through a coarse glass filter, and the collected solid waswashed with isopropanol (64 mL) and methyl tert-butyl ether (64 mL). Theresulting, free-flowing, tan solid was dried in vacuo at 70° C. to give17.4 g (36%) of (R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidinemono-citrate (mixture of Forms II and III) as a tan solid. ¹H NMR (D₂O,400 MHz) δ 8.85 (s, 1H), 8.70 (s, 1H), 6.50 (d, J=17 Hz, 1H), 6.35 (dd,J=7 Hz, J=17 Hz, 1H), 3.43-3.50 (m, 1H), 3.34-3.43 (m, 1H), 3.20-3.30(m, 1H), 3,08-3.19 (m, 1H), 3.00-3.08 (m, 1H), 2.77 (d; J=16 Hz, 2H),2.65 (d, J=16 Hz, 2H), 2.16-2.26 (m, 1H), 1.80-1.92 (m, 1H).

Example 11 Screen for hydrochloric acid addition salts of(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine

(R)-5-((E)-2-Pyrrolidin-3-ylvinyl)pyrimidine free base was dissolved ineither, isopropyl acetate, tetrahydrofuran, methyl isobutyl ketone,acetonitrile, or isopropyl alcohol. The resulting solution was treatedwith 1 eq. of HCl delivered in one of the following forms: 1M in diethylether, 1M in water, 5M in isopropyl alcohol or 4M in dioxane. Themixture was incubated at 50° C./ambient temperature (4 h cycles) for 24h. Where the experiment resulted in a stable solid, the material wasanalyzed by XRPD.

Example 12 Screen for “mono” acid addition salts of(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine

(R)-5-((E)-2-Pyrrolidin-3-ylvinyl)pyrimidine free base (10 mg, 0.057mmol) was dissolved in either isopropyl acetate or acetonitrile. Thesolutions were treated with 1 eq. of the corresponding acid (see below),warmed to 50° C., and cooled slowly to ambient temperature overnight.The solvent was then evaporated under vacuum without heating, and theresidues analyzed by XRPD. The solids are then stored in a humiditychamber at 40° C. and 75% RH for a week, and re-analyzed by XRPD.

In the cases where the experiment did not yield a crystalline solid, thesamples were maturated in tetrahydrofuran and isopropyl alcohol, andwhere a solid was obtained, the solid was analyzed by XRPD and stored inthe humidity chamber for a week to assess stability.

The following acids were screened, using the above procedures forforming “mono” acid addition salts: hydrochloric acid, sulfuric acid,methanesulfonic acid, maleic acid, phosphoric acid,1-hydroxy-2-naphthoic acid, ketoglutaric acid, malonic acid, L-tartaricacid, fumaric acid, citric acid, L-malic acid, hippuric acid, L-lacticacid, benzoic acid, succinic acid, adipic acid, acetic acid, nicotinicacid, propionic acid, orotic acid, 4-hydroxybenzoic acid, anddi-p-Toluoyl-D-tartaric acid.

Example 13 Screen for “hemi” acid addition salts of(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine

(R)-5-((E)-2-Pyrrolidin-3-ylvinyl)pyrimidine free base (10 mg, 0.057mmol) was dissolved in either isopropyl acetate or acetonitrile. Thesolutions were then treated with 0.5 eq. of the corresponding acid (seebelow), warmed to 50° C., and cooled slowly to ambient temperatureovernight. The solvent was then evaporated under vacuum without heating,and the residues analyzed by XRPD. The solids were then stored in thehumidity chamber at 40° C. and 75% RH for a week, and re-analyzed byXRPD.

In the cases where the experiment did not yield a crystalline solid,these samples were maturated in tetrahydrofuran and isopropyl alcohol,and where a solid was obtained, the solid is analyzed by XRPD and storedin the humidity chamber for a week to assess stability.

The following acids were screened, using the above procedures forforming “hemi” acid addition salts: sulfuric acid, maleic acid,phosphoric acid, ketoglutaric acid, malonic acid, L-tartaric acid,fumaric acid, citric acid, L-malic acid, succinic acid, adipic acid, anddi-p-toluoyl-D-tartaric acid.

Example 14 General Scale-Up Procedure for Selected Salts of(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine

A number of (R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine salts werechosen to scale-up to ˜200 mg for further characterization. These saltforms include: citrate (mono and hemi), orotate (mono),4-hydroxybenzoate (mono), di-p-toluoyl-D-tartrate (mono and hemi),maleate (mono and hemi), and fumarate (mono and hemi).

(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine free base (189 mg, 1.077mmol, was dissolved in acetonitrile. The solution was then treated with1.1 eq. of the corresponding acid for the preparation of the mono salt,and 0.5 eq. for the preparation of the hemi salt. The mixture was warmedup to 50° C. and cooled down slowly to ambient temperature overnight.

The solid obtained was filtered and dried under suction before beinganalyzed by XRPD, and ¹H-NMR. TGA experiments were performed todetermine content of water or other solvents, and DSC experiments wererun to establish stability of the isolated forms and the possibility ofnew forms for each salt. DVS experiments were used to assesshygroscopicity of the salts. HPLC purity and thermodynamic solubilitywere also measured for each salt.

Example 15 (R)-5-((E)-2-Pyrrolidin-3-ylvinyl)pyrimidine mono-citrateForm I

(R)-5-((E)-2-Pyrrolidin-3-ylvinyl)pyrimidine mono-citrate Form I wasobtained according to the mono salt screening procedure, from isopropylacetate, by evaporation and maturation in tetrahydrofuran.Alternatively, the mono-citrate Form I was obtained according to themono salt screening procedure, from acetonitrile, by evaporation andmaturation in isopropyl alcohol. The XRPD diffractogram of(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine mono-citrate Form I isshown in FIG. 2.

Example 16 (R)-5-((E)-2-Pyrrolidin-3-ylvinyl)pyrimidine mono-citrateForm II

A suspension of the (R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidinemono-citrate Forms II and Ill mixture in methanol was heated to 50° C.and stirred for 1 h. The suspension was subsequently cooled to 20° C. ata rate of −30° C./h, followed immediately by heating back to 50° C. at arate of +30° C./h. Heating was discontinued upon reaching 50° C., andthe suspension was cooled and stirred at ambient temperature for 16 h.The suspension was filtered, and any residual material in the flask wasrinsed out with additional methanol. The residue was dried at 70° C. invacuo for 16 h to give (R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidinemono-citrate Form II. The XRPD diffractogram of(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine mono-citrate Form II isshown in FIG. 3.

Example 17 Amorphous (R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidinemono-citrate

Amorphous (R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine mono-citrate wasprepared by freeze drying a solution of(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine mono-citrate Form II inwater. The XRPD diffractogram of amorphous(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine mono-citrate is shown inFIG. 1.

Example 18 (R)-5-((E)-2-Pyrrolidin-3-ylvinyl)pyrimidine mono-citrateForm III

(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine mono-citrate Form III wasprepared by allowing amorphous(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine mono-citrate to stand atambient temperature for two hours. The XRPD diffractogram of(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine mono-citrate Form III isshown in FIG. 4.

Example 19 (R)-5-((E)-2-Pyrrolidin-3-ylvinyl)pyrimidine mono-citrateForm IV

(R)-5-((E)-2-Pyrrolidin-3-ylvinyl)pyrimidine mono-citrate Form IV wasobtained by maturation of Form II in acetone/methyl isobutyl ketone. TheXRPD diffractogram of mono-citrate Form IV is shown in FIG. 5.

Example 20 (R)-5-((E)-2-Pyrrolidin-3-ylvinyl)pyrimidinemono-(R)-(−)-orotate salt

Orotic acid (0.965 g, 6.18 mmol) was added as a solid to a stirring, hotsolution of (R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine free base(1.084 g, 6.18 mmol) in 2-propanol (10 mL) in a round-bottomed flask.The resulting mixture of solids was heated under reflux for 5 min,cooled to ambient temperature and stirred overnight. The light-beigepowder was filtered, washed with 2-propanol (10, 8 mL) and dried in avacuum oven (air bleed) at 50° C. for 20 h to give 1.872 g (77.9%) of anoff-white to white, lumpy solid, mp 230-233° C. ¹H NMR (D₂O): δ 8.80 (s,1H), 8.60 (s, 2H), 6.40 (d, 1H), 6.25 (dd, 1H), 5.93 (s, 1 H, ═CH oforotic acid, indicating a mono-salt stoichiometry), 3.38 (dd, 1 H), 3.29(m, 1H), 3.17 (m, 1H), 3.04 (m, 1H), 2.97 (dd, 1H), 2.13 (m, 1H), 1.78(m, 1H). Elemental analysis results suggests the presence of excessorotic acid and a 1:1.1 base:orotic acid salt stoichiometry. ElementalAnalysis: Calculated for C₁₀H₁₃N₃ C₅H₄N₂O₄: (C, 54.38%; H, 5.17%, N,21.14%); Found: (C, 53.49%, 53.44%; H, 5.04%, 5.10%; N, 20.79%, 20.84%).

Example 21 (R)-5-((E)-2-Pyrrolidin-3-ylvinyl)pyrimidine mono-orotateForm I

(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine free base (189 mg, 1.077mmol, freshly prepared) was dissolved in acetonitrile (5 ml). Thesolution was then treated with 1.1 eq. of an orotic acid solution (1 Min ethanol) at ambient temperature. The mixture was warmed up to 50° C.and cooled down slowly to ambient temperature overnight. The solidobtained was filtered and dried under suction before being analysed byXRPD, and ¹H-NMR. The XRPD diffractogram of(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine mono-orotate Form I isshown in FIG. 6.

Example 22 (R)-5-((E)-2-Pyrrolidin-3-ylvinyl)pyrimidine mono-maleateForm I

(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine free base (189 mg, 1.077mmol, freshly prepared) was dissolved in acetonitrile (5 ml). Thesolution was then treated with 1.1 eq. of an maleic acid solution (1M intetrahydrofuran) at ambient temperature. The mixture was warmed up to50° C. and cooled down slowly to ambient temperature overnight. Thesolid obtained was filtered and dried under suction before beinganalysed by XRPD, and ¹H-NMR. The XRPD diffractogram of(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine mono-maleate Form I isshown in FIG. 7.

Example 23 (R)-5-((E)-2-Pyrrolidin-3-ylvinyl)pyrimidine mono-maleateForm II

(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine mono-maleate (Form I) wasslurried in ethanol and incubated at 50° C./r.t. 4 h-cycle for 48 h.XRPD analysis of the solid showed Form II. The XRPD diffractogram of(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine mono-maleate Form II isshown in FIG. 8.

Example 24 (R)-5-((E)-2-Pyrrolidin-3-ylvinyl)pyrimidine mono-oxalate

Oxalic acid (0.516 g, 5.73 mmol) was added as a solid to a stirring,warm solution of (R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine (1.00 g,5.70 mmol) in ethanol (10 mL). The salt precipitated upon furtherwarming of the solution. To facilitate stirring, the mixture was dilutedwith ethanol (6 mL), and the lumps were broken with a spatula. Themixture was cooled to ambient temperature and was left standingovernight. The light-beige powder was filtered, washed with ethanol, anddried in a vacuum oven at 50° C. for 6 h to give 1.40 g (92.3%) of acreamy-white, fluffy powder, mp 149-151° C. ¹H NMR (DMSO-d₆): δ 9.03 (s,1H), 8.86 (s, 2H), 6.56 (m, 2H), 3.40 (dd, 1H), 3.31 (m, 1H), 3.18 (m,1H), 3.08 (m, 1H), 2.96 (dd, 1H), 2.15 (m, 1H), 1.80 (m, 1H), ¹³C NMR(DMSO-d₆): δ 164.90 (C═O of oxalic acid), 156.97, 154.17, 133.66,130.31, 124.20, 48.70, 44.33, 40.98, 30.42. Elemental analysis:Calculated for C₁₀H₁₃N₃·C₂H₂O₄ (C, 54.33%; H, 5.70%, N, 15.84%); Found(C, 54.39%, 54.29%; H, 5.68%, 5.66%; N, 15.68%, 15.66%).

Example 25 (R)-5-((E)-2-Pyrrolidin-3-ylvinyl)pyrimidinehemi-di-p-toluoyl-D-tartarate

Solid di-p-toluoyl-D-tartarate salts was obtained according to the“hemi” salt screening procedure from isopropyl acetate or acetonitrileby evaporation, or by evaporation if isopropyl acetate followed bymaturation with tetrahydrofuran or by evaporation of acetonitrilefollowed by maturation with isopropyl alcohol.

The following procedure was used to make a larger quantity of the salt.(+)-O,O′-Di-p-toluoyl-D-tartaric acid (1.103 g, 2.85mmol) was added as asolid to a stirring, warm solution of(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine free base (1.007 g, 5.74mmol) in ethanol (10 mL). A few insoluble solids precipitated thatfailed to dissolve upon heating the mixture to reflux. The light ambersolution (with a few fine solids) was stirred for 4-5 h and then allowedto stand at ambient temperature overnight. The precipitation of the saltas a light beige powder was slow. After stirring for 15 days, the solidswere filtered, washed with ethanol (5 mL) and dried in a vacuum oven at50° C. for 21 h to give 1.50 g (71.5%) of an off-white to slightlyyellow-tinged powder, mp 178-180° C. ¹H NMR (DMSO-d₆) confirms the 1:0.5base:acid salt stoichiometry. ¹H NMR (DMSO-d₆): δ 10.30 (broad s, ˜1H),9.02 (s, 1H), 8.80 (s, 2H), 7.87 (d, 2H, —C₆H₄-, indicating a hemi-saltstoichiometry), 7.27 (d, 2H, —C₆H₄-, indicating a hemi-saltstoichiometry), 6.40 (dd, 1 H), 6.34 (d, 1H), 5.58 (s, 1H, CH(CO₂H)—O-of acid moiety, indicating a hemi-salt stoichiometry), 3.21 (dd, 1H),3.14 (m, 1H), 3.00 (m, 1H), 2.86 (m, 1H), 2.75 (dd, 1H), 2.30 (s, 3H,—CH₃ of acid moiety, indicating a hemi-salt stoichiometry), 1.93 (m,1H), 1.61 (m, 1H).

Example 26 (R)-5-((E)-2-Pyrrolidin-3-ylvinyl)pyrimidinehemi-di-p-benzoyl-D-tartarate

(+)-O,O′-Di-benzoyl-D-tartaric acid (1.025 g, 2.72 mmol) was added as asolid to a stirring, warm solution of(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine free base (1.003 g, 5.72mmol) in ethanol (10 mL). The mixture was heated to near reflux on a hotplate, producing a light amber solution. The resulting solution wascooled to ambient temperature and was left standing overnight. Becauseno solids were present, the solution was slowly evaporated in a fumehood, affording tan-brown, gummy solids. Isopropyl acetate (10 mL) wasadded and with spatula scraping and stirring, light beige solids aredeposited. The mixture was stirred overnight. The solids were filtered,washed with isopropyl acetate (2×5 mL) and dried in a vacuum oven at 50°C. for 24 h to give 1.93 g (95.2%) of an off-white powder, mp 155-160°C. ¹H NMR (DMSO-d₆) confirmed the 1:0.5 base:acid salt stoichiometry. ¹HNMR (DMSO-d₆): δ 10.25 (broad s, ˜1H), 9.02 (s, 1H), 9.80 (s, 2H), 7.98(d, 2H C₆H₅-), 7.57 (m, 1H, C₆H₅-), 7.48 (m, 2H, C₆H₅—), 6.38 (m, 2H),5.61 (s, 1 H, —CH(CO₂H)—O- of acid moiety, indicating a hemi-saltstoichiometry), 3.22 (dd, 1H), 3.14 (dt, 1H), 3.00 (dt, 1H), 2.88 (m,1H), 2.77 (dd, 1H), 1.92 (m, 1H), 1.61 (m, 1H).

Example 27 (R)-5-((E)-2-Pyrrolidin-3-ylvinyl)pyrimidinehemi-di-p-anisoyl-D-tartarate

(+)-Di-p-anisoyl-D-tartaric acid (1.199 g) was added as a solid to astirring, warm solution of (R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidinefree base (0.999 g) in ethanol (10 mL). The resulting solution, with afew solids present, was stirred and heated in an attempt to dissolve allsolids. The solution became a thick mass. After standing at ambienttemperature for 4-5 h, additional ethanol (10 mL) was added. The mixturecontaining light-beige to cream-colored solids was stirred overnight.The solids were filtered, washed with ethanol (10 mL), and dried in avacuum oven at 50° C. for 21 h to give 1.91 g (87.3%) of a white powder,mp 173-177° C. ¹H NMR (DMSO-d₆) confirmed the 1:0.5 base:acid saltstoichiometry. ¹H NMR (DMSO-d₆): δ 10.20 (broad s, ˜1H), 9.02 (s, 1H),8.80 (s, 2H), 7.93 (d, 2H, —C₆H₄-, indicating a hemi-saltstoichiometry), 7.00 (d, 2H, —C₆H₄-, indicating a hemi-saltstoichiometry), 6.40 (dd, 1 H), 6.34 (d, 1 H), 5.56 (s, 1 H, CH(CO₂H)-O-of acid moiety, indicating a hemi-salt stoichiometry), 3.76 (s, 3H,—OCH₃ of acid moiety, indicating a hemi-salt stoichiometry), 3.22 (dd,1H), 3.14 (m, 1H), 3.01 (m, 1H), 2.85 (m, 1H), 2.75 (m, 1H), 1.92 (m,1H), 1.61 (m, 1H).

Example 28 (R)-5-((E)-2-Pyrrolidin-3-ylvinyl)pyrimidinemono-di-p-toluoyl-D-tartarate

Solid di-p-toluoyl-D-tartarate salts were obtained according to the“mono” salt screening procedure from isopropyl acetate or acetonitrileby evaporation.

The following procedure was used to make a larger quantity of the salt.(+)—O,O′-Di-p-toluoyl-D-tartaric acid (2.205 g, 5.71 mmol) was added asa solid to a stirring, warm solution of(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine free base (1.000 g, 5.70mmol) in ethanol (21 mL). Precipitation of the salt was immediate. Aftergently heating the stirring mixture on a hot plate to near reflux, theresulting mixture was cooled to ambient temperature. The resultingsolids were filtered, washed with ethanol (3×5 mL), and dried in avacuum oven at 50° C. for 13 h to give 3.081 g (96.1%) of a light-beigepowder, mp 181-184° C. ¹H NMR (DMSO-d₆) confirmed the 1:1 saltstoichiometry. ¹H NMR (DMSO-d₆): δ 9.60 (broad s, ˜1 H), 9.03 (s, 1H),8.82 (s, 2H), 7.83 (d, 4H, —C₆H₄-, indicating a mono-saltstoichiometry), 7.27 (d, 4H, —C₆H₄-, indicating a mono-saltstoichiometry), 6.44 (d, 2H), 5.62 (s, 2H, CH(CO₂H)—O- of acid moiety,indicating a mono-salt stoichiometry), 3.30 (dd, 1 H), 3.23 (m, 1 H),3.09 (m, 1 H), 2.95 (m, 1 H), 2.85 (dd, 1 H), 2.33 (6H, —CH₃ of acidmoiety, indicating a mono-salt stoichiometry), 2.02 (m, 1H), 1.69 (m,1H).

Example 29 (R)-5-((E)-2-Pyrrolidin-3-ylvinyl)pyrimidinemono-di-p-benzoyl-D-tartarate

(+)-O,O′-Di-benzoyl-D-tartaric acid (2.05 g, 5.72 mmol) was added as asolid to a stirring, warm solution of(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine free base (0.999 g, 5.69mmol) in ethanol (21 mL) in a round-bottomed flask, producing asolution. After stirring and further heating, precipitation of the saltoccurred in the warm solution. The resulting mixture was cooled toambient temperature over a two-day weekend. The resulting solids werefiltered on a Büchner funnel, washed with ethanol (4×5 mL), and dried ina vacuum oven (air bleed) at 50° C. for 13 h to give 2.832 g (93.0%) ofa light-beige to off-white powder, mp 165-171° C. ¹H NMR (DMSO-d₆)confirmed the 1:1 salt stoichiometry. ¹H NMR (DMSO-d₆): δ 9.65 (broad s,−1H), 9.03 (s, 1H), 9.83 (s, 2H), 7.94 (d, 4H, C₆H₅-), 7.60 (m, 2H,C₆H₅-), 7.50 (m, 4H, C₆H₅-), 6.45 (m, 2H), 5.67 (s, 2H, —CH(CO₂H)—O—ofacid moiety, indicating a mono-salt stoichiometry), 3.31 (dd, 1 H), 3.22(m, 1H), 3.08 (m, 1H), 2.96 (m, 1H), 2.85 (dd, 1H), 2.01 (m, 1H), 1.69(m, 1H).

Example 30 (R)-5-((E)-2-Pyrrolidin-3-ylvinyl)pyrimidinemono-(1S)-10-camphorsulfonate

(1S)-(+)-10-Camphorsulfonic acid (1.329 g, 5.72 mmol) was added as asolid to a stirring, warm solution of(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine free base (1.00 g) in2-propanol (23 mL, 5.70 mmol) in a round-bottomed flask. Upon cooling toambient temperature, there was no precipitation of solids. The solutionwas allowed to stand overnight. Gelatinous material containing whitesolids was observed. After stirring two days, the mixture was dilutedwith 2-propanol (10.5 mL) because stirring this jelly-like white masswas difficult. After overnight stirring, the resulting white powder wasfiltered on a Büchner funnel, washed with 2-propanol (5 mL) (NOTE: Thesolids appeared to have some solubility in 2-propanol) and dried in avacuum oven (air bleed) at 50° C. for 6 h to give 1.47 g (63.2%) of awhite powder, mp 172-173° C. ¹H NMR (DMSO-d₆) confirms the 1:1 saltstoichiometry. After standing seven days, a second crop of light-beigeneedles was observed in the crystallization liquors. This material wasfiltered, washed with 2-propanol (10 mL) and dried in a vacuum oven (airbleed) at 50° C. for 21 h to give 0.245 g of light-beige needles, mp173-174° C. ¹H NMR (DMSO-d₆): δ 9.03 (s, 1 H), 8.87 (s, 2H), 6.57 (m,2H), 3.41 (dd, 1 H) 3.33 (m, 1 H, partially masked by H₂O), 3.21 (m, 1H), 3.10 (m, 1 H), 2.98 (dd, 1 H), 2.89 (d, 1 H, —CH₂- of acid moiety,indicating a mono-salt stoichiometry), 2.64 (m, 1 H), 2.41 (d, 1H, —CH₂-of acid moiety, indicating a mono-salt stoichiometry), 2.25 (t, 0.5 H),2.20 (t 0.5 H), 2.15 (m, 1H), 1.93 (t, 1H), 1.82 (m, 3H), 1.28 (m, 2H,—CH₂- of acid moiety, indicating a mono-salt stoichiometry), 1.03 (s,3H, —CH₃ of acid moiety, indicating a mono-salt stoichiometry), 0.73 (s,3H, —CH₃ of acid moiety, indicating a mono-salt stoichiometry).

Example 31 (R)-5-((E)-2-Pyrrolidin-3-ylvinyl)pyrimidine mono-(1R,2S)-(+)-Camphorate

(1 R,2S)-(+)-Camphoric acid (1.149 g, 5.74 mmol) was added as a solid toa stirring, warm solution of(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine free base (1.00 g, 5.70mmol) in ethanol (14 mL) in a round-bottomed flask. Upon heating, allsolids dissolved, affording a yellow solution. No precipitate forms uponstanding at ambient temperature overnight. The solution was concentratedvia rotary evaporation to an amber-brown foam that was dried undervacuum at 50° C. (air bleed) for 6 h to give 2.098 g of a viscous, amberoil. Isopropyl acetate (10 mL) was added, and the solution was allowedto stand at ambient temperature overnight. There was some evidence ofcrystal nucleation in the gummy, red-amber oil. More isopropyl acetate(10 mL) and 2-propanol (20 drops) was added, and the mixture was gentlyheated and stirred over 48 h. The resulting milky, creamy solids withsome orange lumps were broken with a spatula, and the mixture (colorlessliquor) was stirred overnight. The off-white solids were filtered on aBüchner funnel, washed with cold isopropyl acetate (10 mL) and dried ina vacuum oven (air bleed) at 50° C. for 21 h to give 2.034 g (94.9%) ofan off-white to cream colored powder, mp 157-159° C. ¹H NMR (DMSO-d₆)confirmed the 1:1 salt stoichiometry. ¹H NMR (DMSO-d₆): δ 9.00 (s, 1H),8.85 (s, 2H), 6.58 (dd, 1H), 6.47 (d, 1H), 3.17 (dd, 1H), 3.08 (m, 1H),2.97 (m, 1H), 2.92 (dd, 1H) 2.74 (dd, 1H), 2.61 (dd, 1H), 2.30 (sextet,1H), 2.00 (m, 2H), 1.65 (m, 2H), 1.32 (m, 1H), 1.15 (s, 3H, —CH₃ of acidmoiety, indicating a mono-salt stoichiometry), 1.07 (s, 3H, —CH₃ of acidmoiety, indicating a mono-salt stoichiometry), 0.75 (s, 3H, —CH₃ of acidmoiety, indicating a mono-salt stoichiometry).

Example 32 (R)-5-((E)-2-Pyrrolidin-3-ylvinyl)pyrimidinemono-di-p-anisoyl-D-tartarate

(+)-Di-p-anisoyl-D-tartaric acid (2.388 g, 5.71 mmol) was added as asolid to a stirring, warm solution of(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine free base (1.008 g, 5.75mmol) in ethanol (22 mL) in a round-bottomed flask. Precipitation of thesalt occurred before all of the (+)-di-p-anisoyl-D-tartaric acid hadbeen added. The salt did not dissolve upon heating, but the appearanceof the solids changed, with conversion to a light, fluffy, white powder.The mixture was cooled to ambient temperature and was stirred over 48 h.The resulting solids were filtered on a Büchner funnel, washed withethanol (5×5 mL) and dried in a vacuum oven (air bleed) at 50° C. for 13h to give 3.20 g (94.4%) of an off-white to white, chalky powder, mp173-176° C. ¹H NMR (DMSO-d₆) confirms the 1:1 salt stoichiometry. ¹H NMR(DMSO-d₆): δ 9.65 (broad s, −1 H), 9.03 (s, 1H), 8.82 (s, 2H), 7.89 (d,4H, —C₆H₄—, indicating a mono-salt stoichiometry), 7.01 (d, 4H, —C₆H₄—,indicating a mono-salt stoichiometry), 6.44 (m, 2H), 5.60 (s, 2H,CH(CO₂H)-O- of acid moiety, indicating a mono-salt stoichiometry), 3.79(s, 6H, —OCH₃ of acid moiety, indicating a mono-salt stoichiometry),3.30 (dd, 1H), 3.22 (m, 1H), 3.09 (m, 1H), 2.95 (m, 1H), 2.84 (m, 1H),2.01 (m, 1H), 1.69 (m, 1H).

Example 33 (R)-5-((E)-2-Pyrrolidin-3-ylvinyl)pyrimidinemono-(R)-(−)-Phencyphos salt

(R)-(−)-Phencyphos (1.391 g, 5.77 mmol) was added as a solid to astirring solution of (R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine freebase (1.006 g, 5.73 mmol) in ethanol (10 mL) in a round-bottomed flask.Most of the solids dissolved upon stirring at ambient temperature, andall solids dissolved with gentle heating. The stirring, amber solutionwas heated to reflux, cooled to ambient temperature and was allowed tostand overnight. The resulting white, needle-like crystals were filteredon a Büchner funnel, washed with cold ethanol (5 mL) and dried in avacuum oven (air bleed) at 50° C. for 18 h to give 0.811 g (33.9%) ofoff-white crystals, mp 197-201° C. ¹H NMR (DMSO-d₆) confirms the 1:1salt stoichiometry. ¹H NMR (DMSO-d₆): δ 9.81 (broad s, −1 H), 9.02 (s, 1H), 8.85 (s, 2H), 7.27 (m, 5H, C₆H₅—), 6.56 (dd, 1H), 6.48 (d, 1H), 5.00(d, 1H, —O—CH— of acid moiety, indicating a mono-salt stoichiometry),4.00 (d, 1H, —O—CH₂- of acid moiety, indicating a mono-saltstoichiometry), 3.48 (dd, 1H, —O—CH₂- of acid moiety, indicating amono-salt stoichiometry), 3.36 (dd, 1H), 3.30 (m, 1H), 3.17 (m, 1H),3.07 (m, 1H), 2.93 (dd, 1H), 2.12 (m, 1H), 1.78 (m, 1H), 0.79 (s, 3H,—CH₃ of acid moiety, indicating a mono-salt stoichiometry), 0.60 (s, 3H,-CH₃ of acid moiety, indicating a mono-salt stoichiometry).

Although specific embodiments of the present invention are hereinillustrated and described in detail, the invention is not limitedthereto. The above detailed descriptions are provided as exemplary ofthe present invention and should not be construed as constituting anylimitation of the invention. Modifications will be obvious to thoseskilled in the art, and all modifications that do not depart from thespirit of the invention are intended to be included with the scope ofthe appended claims.

We claim:
 1. A mono-citrate salt of a compound(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine substantially free of(S)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine.
 2. A pharmaceuticalcomposition comprising (R)-5-(((E)-2-pyrrolidin-3-ylvinyl)pyrimidinemono-citrate substantially free of(S)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine mono-citrate and one ormore pharmaceutically acceptable carriers, diluents, excipients, oradjuvants.
 3. A method of treating constipation in a patient in needthereof comprising administration of(R)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine mono-citrate substantiallyfree of (S)-5-((E)-2-pyrrolidin-3-ylvinyl)pyrimidine mono-citrate.